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Endocrinology Vol. 141, No. 7 2370-2376
Copyright © 2000 by The Endocrine Society

ARTICLES

Activation of Signal Transducer and Activator of Transcription-3 during Proliferative Phases of 3T3-L1 Adipogenesis1

Jianbei Deng, Kunjie Hua, Steven S. Lesser and Joyce B. Harp

Department of Nutrition, University of North Carolina, Chapel Hill, North Carolina 27599

Address all correspondence and requests for reprints to: Joyce B. Harp, M.D., Department of Nutrition, University of North Carolina, CB# 400 McGavran-Greenberg Hall, Chapel Hill, North Carolina 27599. E-mail: jharp{at}sph.unc.edu


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
Signal transducer and activator of transcription-3 (STAT3) is abundantly expressed in preadipocytes and adipocytes, but little is known about its activation status or functional role during adipogenesis. In this report we investigate STAT3 activation in 3T3-L1 preadipocytes before and after differentiation into adipocytes. STAT3 was highly tyrosine phosphorylated and bound to DNA in proliferating preadipocytes, but not in growth-arrested preadipocytes or adipocytes. In growth-arrested confluent preadipocytes, induction of differentiation with methylisobutylxanthine, dexamethasone, and high dose insulin led to a delayed, but prolonged (3-day), increase in STAT3 tyrosine phosphorylation. This increase in STAT3 phosphorylation coincided temporally with postconfluent preadipocyte mitotic clonal expansion. Insulin and methylisobutylxanthine alone, but not dexamethasone, induced STAT3 tyrosine phosphorylation in postconfluent cells. Diminution of endogenous STAT3 expression by antisense morpholino oligonucleotides significantly decreased preconfluent preadipocyte proliferation. Collectively, these findings suggest a regulatory role for STAT3 during the proliferative phases of adipogenesis.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
IN WHITE adipose tissue, preadipocytes exist in close proximity to adipocytes and respond to positive energy balance by proliferating, then differentiating into adipocytes (1, 2). Mature adipocytes then accommodate excess energy through enhanced triacylglycerol storage. Although much is known about the regulatory events surrounding preadipocyte differentiation and adipocyte lipogenesis, less is known about earlier events of preadipocyte replication. Many of the advances in our understanding of adipogenesis are based on studies in murine 3T3-L1 cells. 3T3-L1 preadipocytes are committed fibroblast-like cells that replicate in culture until they reach confluence (3). At confluence, the preadipocytes undergo cell-cell contact-inhibited growth arrest. Upon stimulation with high dose insulin, dexamethasone, and methylisobutylxanthine (MIX) for 2 days, preadipocytes undergo several rounds of mitotic clonal expansion, then exit the cell cycle and begin to express adipocyte-specific genes (4). Approximately 5 days after differentiation, more than 90% of the cells display the characteristic lipid-filled adipocyte phenotype. The induction of differentiation and subsequent expression of adipocyte-specific genes depend on transcriptional activation and expression of at least two families of transcription factors; the CCAAT/enhancer-binding proteins (C/EBP) {alpha}, ß, {delta}, and peroxisome proliferator-activated receptor {gamma}2 (PPAR{gamma}2) (4). C/EBP{alpha} plays a dual role in acquisition of the growth-arrested state and initiation of the adipocyte phenotype. PPAR{gamma}2 synergizes with C/EBP{alpha} to promote adipocyte differentiation (5). Another group of transcription factors, signal transducers and activators of transcription (STATs), was recently identified in 3T3-L1 cells, but their role in adipogenesis has not been defined (6).

In mammalian cells, STAT1, -2, -3, -4, -5A, -5B, and -6 comprise a group of latent cytoplasmic transcription factors that are activated by cytokines, peptides, and growth factors (7). Activated receptor and nonreceptor tyrosine kinases phosphorylate STATs on critical tyrosine residues in the carboxyl-terminal domain. STATs then homo- or heterodimerize through reciprocal phosphotyrosine-SH2 domain interaction and translocate to the nucleus, where they bind to specific DNA regulatory sequences to stimulate the transcription of targeted effector genes. STATs become latent again upon dephosphorylation and translocation back to the cytoplasm (8).

Studies in 3T3-L1 cells have shown that STAT1, STAT3, STAT5, and STAT6 are expressed in 3T3-L1 preadipocytes and adipocytes, but expression levels change during the differentiation program (6). The expression of STAT1 and STAT5 increases significantly after the induction of differentiation, STAT3 levels increase slightly, and STAT6 expression does not change. Although a number of activating ligands have been identified for the various STATs in preadipocytes and adipocytes, a clear picture of their functional role has not been defined. For STAT3, leukemia inhibitory factor, interferon-{gamma}, and oncostatin M induced STAT3 tyrosine phosphorylation and nuclear translocation in 3T3-L1 adipocytes (9, 10). Other traditional activators of STAT3, interleukin-6 (IL-6) and platelet-derived growth factor, induced STAT3 tyrosine phosphorylation to a lesser extent. Epidermal growth factor had no effect on STAT3 activation. In human preadipocytes, IL-6, IL-11, leukemia inhibitory factor, and oncostatin M activated STAT3, which then stimulated transcription of the P450 aromatase gene through an interferon-{gamma} active site element in the I.4 promoter (11). Aromatase induces estrogen biosynthesis from androstenedione in adipose tissue.

Because STAT3 promotes the proliferation of other mescenchymal cells, i.e. vascular smooth muscle cells (12), we hypothesized that STAT3 may play a critical role during proliferative stages of 3T3-L1 adipogenesis. In this report we provide evidence that STAT3 is highly activated in proliferating preconfluent and postconfluent preadipocytes, but not in growth-arrested postconfluent preadipocytes or differentiated adipocytes. Decreased STAT3 expression by antisense oligonucleotides in preconfluent preadipocytes led to a decrease in cell proliferation. STAT3 activation during preconfluent preadipocyte proliferation occurred in the absence of exogenous ligands, whereas insulin and MIX mediated STAT3 activation in postconfluent cells.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Materials
3T3-L1 cells were obtained from American Type Culture Collection(Rockville, MD). Affinity-purified rabbit polyclonal phosphotyrosine 705-specific anti-STAT3 (STAT3-PY705), rabbit polyclonal anti-STAT3, and anti-STAT1-PY701 antibodies were purchased from New England Biolabs, Inc. (Beverly, MA). Monoclonal antibodies to the C-terminal region of STAT1 were obtained from Transduction Laboratories (Lexington, KY). Monoclonal STAT3 (H190) TransCruz supershift antibodies, anti-STAT6 antibodies (S20), and protein A/G plus agarose were obtained from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA). STAT3 gel shift oligonucleotides were synthesized at the University of North Carolina Lineberger Comprehensive Cancer Center Nucleic Acid Core Facility. The enhanced chemiluminescence detection kit, horseradish peroxidase-conjugated secondary antibodies, and [{alpha}-32P]deoxy-CTP were obtained from Amersham Pharmacia Biotech (Piscataway, NJ). Control (5'-CCTCTTACCTCAGTTACAATTTATA-3') and STAT3 antisense (5'-TGGTTCCACTGAGCCATCCTGCTTGC-3') morpholinos were obtained from Gene Tools, Inc. (Corvallis, OR).

Cell culture
3T3-L1 preadipocytes were cultured in DMEM containing 10% (vol/vol) FBS, 10 mg/ml streptomycin, 100 U/ml penicillin, and 1 mM pyruvate at 37 C in 5% CO2-air. Preadipocytes were studied when preconfluent, confluent, and after differentiation into adipocytes. To induce differentiation, 2-day confluent cells were treated with 0.5 µM dexamethasone, 0.5 mM MIX, and 10 µg/ml insulin in DMEM/10% FBS for 48 h followed by maintenance in DMEM/10% FBS with added insulin for an additional 48 h, then cells were placed back in DMEM/10% FBS.

For loading of morpholinos, 3T3-L1 cells were plated at a density of 1 x 105 cells/35-mm dish for 24 h. Morpholinos (3 µM) were added by the scrape-loading method (13). Briefly, morpholinos were added to the plated cells, then preadipocytes were immediately scraped with a rubber policemen. Cells were then removed and replated in toto into new 6-well plates or in 96-well plates at a density of 5 x 104 cells/well for measurement of cell proliferation. Cell proliferation was measured by the Nonradioactive MTS Cell Proliferation kit (Promega Corp., Madison, WI) as directed by the supplier.

Immunoblot analysis
3T3-L1 cells were washed twice in PBS with 1 mM orthovanadate, then placed immediately in sample buffer [1% Nonidet P-40, 20 mM Tris-HCl (pH 8.0), 150 mM NaCl, 1 mM EDTA, 0.1% NaN3, 10 µg/ml aprotinin, 1 µM pepstatin, 16.4 µg/ml leupeptin, 1 mM phenylmethylsulfonylfluoride, 0.1 mM Na3VO4, 2% SDS, and 10% glycerol] without dithiothreitol or tracking dye. Lysates were heated, and protein concentrations were determined before adding 100 mM dithiothreitol and tracking dye. Protein concentrations were determined in cell lysates using the Bio-Rad Laboratories, Inc., DC protein determination kit (Richmond, CA). BSA was used as standard. Samples were heated for 5 min at 95 C, separated by 8–10% SDS-PAGE, and analyzed by immunoblotting as previously described (12, 14). Immunoblots were developed with the enhanced chemiluminescence kit.

Electrophoretic mobility shift assay
Whole cell extracts were prepared as previously described and stored at -80 C (15). The STAT3 consensus binding motif corresponding to the high affinity sis-inducing element (hSIE) oligonucleotide (5'-GTCGACATTTCCCGTAAATCGTCGA-3') was used as a probe for gel shift assays (16). The binding reaction mixture containing approximately 20,000 cpm labeled DNA, 2.5 µg poly(dI-dC), and 10 µg protein in 40 mM KCl, 1 mM MgCl2, 0.1 mM EDTA, 20 mM HEPES (pH 7.9), 6% glycerol, and 0.5 mM DTT was incubated on ice for 20 min. Where indicated, 400 x unlabeled probe or 1 µg anti-STAT3 antibody was added to the binding reaction. The reactions were analyzed by 4.5% PAGE in 0.5 x Tris-glycine, then dried and visualized by autoradiography.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
STAT3 is active in 3T3-L1 preadipocytes, but not in adipocytes
To determine STAT3 activation status during adipogenesis, we performed Western blot analysis with phosphotyrosine-specific STAT3 antibodies (STAT3-PY705) in 3T3-L1 preadipocytes at multiple times before and after differentiation into adipocytes (Fig. 1AGo). STAT3 tyrosine phosphorylation increased as cells proliferated and became confluent, then decreased sharply to barely detectable levels 2 days after cells reached confluence. There was a transient increase in STAT3 phosphorylation 24 h after differentiation, which then declined and remained low in differentiating preadipocytes and mature adipocytes. Although there were dramatic changes in STAT3 tyrosine phosphorylation, STAT3 protein levels did not change during proliferation and differentiation. To test whether this pattern of tyrosine phosphorylation was specific for STAT3 or representative of all STATs, we performed similar studies with phosphotyrosine-specific STAT1 91/84 antibodies. Tyrosine phosphorylation of STAT1 was barely detectable in proliferating preadipocytes and did not become activated until 4 days after differentiation, a time when the cells began to appear as adipocytes (Fig. 1BGo). Low levels of STAT1 p91 expression occurred in proliferating preadipocytes (P-3), but STAT1 p84 was not detectable. With differentiation, there was a transient decrease in STAT1 p91 expression, followed by an increase in p91/84 expression with adipocyte formation. STAT6 expression remained constant throughout the differentiation program, except for a slight decrease 6 days after differentiation (Fig. 1CGo). STAT6 was not tyrosine phosphorylated on residue 641 throughout the program of adipogenesis (data not shown).



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  		Figure 1. STAT3 is tyrosine phosphorylated during 3T3-L1 preadipocyte proliferation. Lysates were prepared from 3T3-L1 cells during progressive stages of preadipocyte proliferation and differentiation. P-3, P-2, and P-1 denote preadipocytes recovered 3, 2, and 1 day(s) before MDI stimulation. A1 to A8, Cells 1–8 days after differentiation into adipocytes. A, Western blot analysis (IB) of 100 µg protein was performed with anti-STAT3-PY. Blots were stripped and preprobed with anti-STAT3 antibodies (A), with anti-STAT1-PY and anti-STAT1 antibodies (B), or with anti-STAT6 (S20) antibodies (C). Results are representative of at least three separate experiments.

 
STAT3 tyrosine phosphorylation induces dimerization, nuclear translocation, and binding to specific sequences in the promoter regions of responsive genes (7). To verify whether STAT3 tyrosine phosphorylation observed in Western blots was also associated with DNA binding, electrophoretic mobility gel shift assays were performed using a hSIE probe and whole cell extracts from preadipocytes and adipocytes. Figure 2AGo shows that STAT3 DNA binding was high in proliferating preadipocytes (P-2) and low in fully differentiated adipocytes (A8), a result that correlates well with the phosphotyrosine data we collected. The STAT3-DNA complex in preadipocytes was supershifted by the addition of an anti-STAT3 antibody, indicating that the complex contained STAT3 protein. To determine whether the changes in STAT3 tyrosine phosphorylation seen immediately before and after differentiation were reflected in STAT3 DNA binding, gel shift analysis was performed in cell extracts 2 days before, just before (day 0), and 1 day after MIX-dexamethasone-insulin (MDI) stimulation. The pattern of STAT3 DNA binding followed that of tyrosine phosphorylation of STAT3 (Fig. 2BGo). However, there was still detectable DNA binding on day 0, when STAT3 tyrosine phosphorylation was barely detectable.



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  		Figure 2. STAT3 binds DNA in untreated 3T3-L1 preadipocytes, but not in untreated adipocytes. Whole cell extracts (10 µg) were prepared (A) from proliferating 3T3-L1 preadipocytes 2 days before confluence (P-2) and from adipocytes 8 days after differentiation (A8) or B) from 3T3-L1 preadipocytes 2 days before confluence (P-2), 2 days after confluence (0), and 1 day after differentiation (A1). Electrophoretic mobility shift assays were performed with a radiolabeled hSIE oligonucleotide probe. Supershift of protein-DNA complexes was performed with 1 µg anti-STAT3 (H190) TransCruz gel shift antibody (Santa Cruz Biotechnology, Inc.). Cold competitor DNA was added at 400 x labeled probe. Results are representative of one or two separate experiments.

 
STAT3 regulates preadipocyte proliferation
To elucidate a potential role for STAT3 during adipogenesis, preconfluent preadipocytes were loaded with antisense morpholino oligonucleotides to the translational start site of STAT3 or control morpholinos containing a random sequence. Treatment of cells (24 h after plating) with STAT3 antisense morpholino oligonucleotides led to a decrease in STAT3 protein expression at 24 and 48 h. Levels of STAT3 began to rise at 72 h, but remained lower (40–60%) than those in cells loaded with control morpholinos (Fig. 3Go, A and B). We did not detect any difference in tyrosine-phosphorylated STAT3 between control and antisense-treated cells, but levels were barely detectable in both cell treatments until 72 h after antisense loading. When the same blots were stripped and reprobed with anti-STAT6, there was no difference in STAT6 expression between antisense-loaded and control cells at all time points.



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  		Figure 3. Morpholino antisense treatment targeting STAT3 decreases STAT3 expression. 3T3-L1 preadipocytes were scrape-loaded with 3 µM antisense STAT3 morpholinos (AS) or control morpholino oligonucleotides (C) 1 day after plating. A, Cell lysates (40 µg) were recovered 24, 48, and 72 h later for Western blot analysis with anti-STAT3 antibodies, then stripped and reprobed with anti-STAT6 antibodies or anti-STAT3-PY antibodies. B, Densitometric analysis was performed on three separate blots, and antisense values are expressed as a percentage of control oligonucleotide values obtained at the same time. The values at 24 and 48 h represent the mean ± SD of three separate experiments. The 72 h point is an average of two separate experiments.

 
As STAT3 activation was coincident with proliferative phases of adipogenesis, we hypothesized that blocking STAT3 expression would lead to a decrease in preadipocyte proliferation. 3T3-L1 cells plated for 24 h were loaded with morpholinos, and cell proliferation was measured over time. In cells loaded with antisense oligonucleotides, there was a significant (P < 0.01) decrease in preadipocyte proliferation, measured by the MTS assay, relative to that in control loaded cells (Fig. 4Go). The degree of inhibition of cell proliferation was most pronounced at times when STAT3 expression was lowest (24 and 48 h). As STAT3 expression increased, proliferation also increased. There was no detectable difference in cytotoxicity measured by lactate dehydrogenase release between control and antisense-treated cells (data not shown).



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  		Figure 4. Morpholino antisense treatment targeting STAT3 decreases 3T3-L1 proliferation. 3T3-L1 preadipocyte proliferation was measured by the MTS assay in cells loaded with STAT3 antisense and control morpholino oligonucleotides 18, 24, 48, 72, and 115 h later (see Materials and Methods). Data are presented as the percent increase over values obtained 24 h after plating and just before loading with morpholinos. Values are the mean ± SD of three separate experiments. *, P < 0.01 when the control group was compared with the antisense group for each time point by t test.

 
Earlier studies suggested that MDI stimulation induced STAT3 tyrosine phosphorylation in postconfluent preadipocytes (Fig. 1AGo). This transient increase in STAT3 activation appeared to coincide with MDI-induced mitotic clonal expansion. Thus, we next measured STAT3 tyrosine phosphorylation and cell proliferation simultaneously at multiple times after the induction of differentiation in 2-day postconfluent preadipocytes. STAT3-PY705 increased 6 h after MDI stimulation and peaked between 0.5–1.5 days after MDI stimulation (Fig. 5AGo). STAT3 phosphorylation remained above baseline for as long as 3 days. This increase in MDI-induced STAT3 tyrosine phosphorylation coincided temporally with the induction of mitotic clonal expansion (Fig. 5BGo).



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  		Figure 5. STAT3 tyrosine phosphorylation correlates with postconfluent preadipocyte mitotic clonal expansion. STAT3 tyrosine phosphorylation measured by Western blot analysis (A) and cell proliferation measured by the MTS assay (B) were simultaneously assayed in postconfluent preadipocytes immediately before and after MDI differentiation at the indicated time points. Results are representative of two separate experiments.

 
Insulin and MIX, but not dexamethasone, induce STAT3 tyrosine phosphorylation
To determine which component of MDI was responsible for STAT3 activation, postconfluent preadipocytes were stimulated with each agent alone for 24 h or in combination with one other agent before recovering lysates for Western blot analysis. Figure 6Go shows that the addition of insulin or MIX alone was sufficient to induce tyrosine phosphorylation of STAT3. Treatment with insulin and MIX combined produced the greatest stimulatory effect. Dexamethasone alone had no effect and completely inhibited both insulin- and MIX-mediated STAT3 tyrosine phosphorylation.



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  		Figure 6. Insulin and MIX, but not dexamethasone, induced STAT3 tyrosine phosphorylation. Postconfluent 3T3-L1 preadipocytes were stimulated with MIX (M), dexamethasone (D), and insulin (I) combined or with each agent alone for 24 h. Cell lysates (50 µg) were assayed by Western blot analysis with anti-STAT3-PY antibody. Blots were stripped and reprobed with anti-STAT3 antibody. Results are representative of four separate experiments.

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
Over the last three decades, 3T3-L1 cells have been studied extensively to understand mechanisms of white adipogenesis. However, most of the work has focused on mechanisms of adipocyte formation rather than preadipocyte proliferation (4). In this study we demonstrated that STAT3 was highly tyrosine phosphorylated and highly bound DNA in proliferating 3T3-L1 preadipocytes, but not in growth-arrested preadipocytes or terminally differentiated adipocytes. At this time, it is unclear whether or not the low levels of STAT3 tyrosine phosphorylation and DNA binding in 3T3-L1 adipocytes are transcriptionally active. Overall, this pattern of activation during adipogenesis was specific for STAT3 and did not occur with STAT1, STAT5, or STAT6 (data not shown for STAT5 and STAT6). The fact that STAT1 was not activated during proliferation is particularly relevant, because STAT1 and STAT3 form heterodimers and are activated by common ligands, i.e. epidermal growth factor and IL-6, in other cell types (17, 18, 19).

Our finding that inhibition of endogenous STAT3 expression with antisense morpholino oligonucleotides decreased preconfluent preadipocyte proliferation indicates that STAT3 activation is upstream, not downstream, of cell proliferation during this stage of adipogenesis. However, in postconfluent preadipocytes the data linking STAT3 activation and cell proliferation are correlative. The exclusive high level activation of STAT3 during proliferation, and not differentiation, is important because another traditional mitogenic signaling pathway, the mitogen-activated protein kinase pathway, showed no difference in activity between 3T3-L1 preadipocytes and adipocytes (20). Our conclusion that STAT3 is critical for preconfluent preadipocyte proliferation is consistent with other studies demonstrating that STAT3 is a regulator of normal and transformed cell proliferation. In rat aortic smooth muscle cells, the vascular mitogen angiotensin II induced STAT3 phosphorylation (12). Inhibition of STAT3 activation through JAK2 inhibitors or electroporated STAT3 blocking antibodies decreased angiotensin II-induced cell proliferation. In transformed squamous epithelial cells derived from human tumors, STAT1 and STAT3 were constitutively active, but cell proliferation was dependent only on STAT3 activation (21). More recently, it was reported that constitutively active STAT3 is an oncogene (22). Expression of constitutively active STAT3 containing substitutions of two cysteine residues in the C-terminal loop of the SH2 domain caused immortalized fibroblasts to undergo cellular transformation and tumor formation in nude mice. This activated STAT3 alone also induced the expression of a number of cell cycle genes. Based on these studies in the literature linking STAT3 to cell proliferation, it was not surprising to find that STAT3 was highly activated during preadipocyte proliferation and not during later stages of differentiation when cells no longer were able to proliferate.

Finally, we found that at the induction of differentiation, insulin and MIX induced STAT3 tyrosine phosphorylation, whereas dexamethasone inhibited it. Others have reported that in 3T3-L1 adipocytes, insulin induced serine phosphorylation of STAT3 on residue 727 and did not induce tyrosine phosphorylation on residue 705 or STAT3 nuclear translocation (10, 23). These results are opposite the findings of another group that showed insulin-induced DNA binding of STAT3 (24). These differences might be explained by the different methods used for assessing STAT3 activation, i.e. phosphoamino acid analysis, nuclear translocation, and gel shift analysis. Our observation that insulin induced the tyrosine phosphorylation of STAT3 on residue 705 differs from previous studies in that we examined 3T3-L1 cells while they were still in the preadipocyte stage. MIX elevates intracellular cAMP by inhibition of the cAMP phosphodiesterase. We know of no other studies linking cAMP to STAT3 activation in any cell, but one study found the opposite effect. In mononuclear blood cells, cAMP inhibited IL-6-induced STAT3 activation (19). In our studies the combined effect of insulin and MIX appeared synergistic in stimulating STAT3 tyrosine phosphorylation. This suggests that the two ligands mediate their effects through distinct pathways or that the concentrations used for each ligand produced a submaximal response of the same pathway. It is also possible that insulin and MIX mediated STAT3 tyrosine phosphorylation through an indirect pathway, i.e. through the synthesis of STAT3-activating ligands, as activation was delayed by 3 h (data not shown). Dexamethasone, insulin, and MIX work in concert to mediate preadipocyte differentiation (4). Our finding that dexamethasone had an effect the opposite of that of insulin and MIX on STAT3 tyrosine phosphorylation is novel. The functional significance of the differential activation of STAT3 by the various ligands is unclear from these studies. Further studies are needed to understand the functional role of STAT3 during preadipocyte differentiation.

In summary, this is the first study to investigate STAT3 activation during proliferative stages of adipogenesis. We provide evidence that STAT3 plays a regulatory role in 3T3-L1 preadipocyte proliferation rather than in the maintenance of the adipocyte phenotype. The mechanism of STAT3 activation appears to be regulated both endogenously and in response to exogenous regulators of preadipocyte differentiation. Future studies are needed to determine the role of STAT3 in adipogenesis in vivo and in the pathogenesis of obesity.


   Footnotes
 
1 This work was supported by USPHS Grant DK-53398, the University Research Council, and the Institute of Nutrition at the University of North Carolina. Back

Received November 4, 1999.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

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F. Machinal-Quelin, M. N. Dieudonne, M. C. Leneveu, R. Pecquery, and Y. Giudicelli
Proadipogenic effect of leptin on rat preadipocytes in vitro: activation of MAPK and STAT3 signaling pathways
Am J Physiol Cell Physiol, April 1, 2002; 282(4): C853 - C863.
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A. W. Harmon and J. B. Harp
Differential effects of flavonoids on 3T3-L1 adipogenesis and lipolysis
Am J Physiol Cell Physiol, April 1, 2001; 280(4): C807 - C813.
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R. L. Mynatt and J. M. Stephens
Agouti regulates adipocyte transcription factors
Am J Physiol Cell Physiol, April 1, 2001; 280(4): C954 - C961.
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K. J. Waite, Z. E. Floyd, P. Arbour-Reily, and J. M. Stephens
Interferon-gamma -induced Regulation of Peroxisome Proliferator-activated Receptor gamma and STATs in Adipocytes
J. Biol. Chem., March 2, 2001; 276(10): 7062 - 7068.
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