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At least 19 recordsLinked to original sources

PubMed1998

Autocrine secretion of TGF-beta 1 and TGF-beta 2 by pre-adipocytes and adipocytes: a potent negative regulator of adipocyte differentiation and proliferation of mammary carcinoma cells.

We have developed an in vitro system to examine the influence of adipocytes, a major mammary stromal cell type, on the growth of a murine mammary carcinoma, SP1. Previously, we have shown that 3T3-L1 adipocytes release a mitogenic factor, hepatocyte growth factor, which strongly stimulates proliferation of SP1 cells. We now show that 3T3-L1 pre-adipocytes secrete active inhibitory molecules which inhibit DNA synthesis in SP1 cells. In addition, latent inhibitory activity is present in conditioned media (CM) from both pre-adipocytes and adipocytes, and is activated following acid treatment. CM also inhibited DNA synthesis in Mv1Lu wild type epithelial cells, but not DR27 mutant epithelial cells which lack TGF-beta type II receptor. Inhibitory activity of CMs was partially abrogated by neutralizing anti-TGF-beta1 and anti-TGF-beta2 antibodies, and was removed following ultrafiltration through membranes of 10,000 Mr but not 30,000 Mr pore size. These results show that the inhibitory effect on DNA synthesis is mediated by TGF-beta1-like and TGF-beta2-like molecules. In addition, acid-treated CM as well as purified TGF-beta inhibited differentiation of pre-adipocytes. Untreated pre-adipocyte CM, but not mature adipocyte CM, spontaneously inhibited adipocyte differentiation. Together, these findings indicate that pre-adipocytes spontaneously activate their own secreted TGF-beta, whereas mature adipocytes do not, and suggest that activation of TGF-beta has a potent negative regulatory effect on adipocyte differentiation and tumor growth. Thus, TGF-beta may be an important modulator of tumor growth and adipocyte differentiation via both paracrine and autocrine mechanisms. These findings emphasize the importance of adipocyte-tumor interactions in the regulation of tumor microenvironment.

3T3 Cells↗

PubMed1986

A thermodynamic approach to hormone-receptor interaction; application to insulin binding to adipocytes, adipocyte plasma membranes and liposomes incorporating adipocyte insulin receptors.

The binding of insulin to its receptor in rat adipocyte and isolated plasma membranes has been measured. The adipocyte insulin receptor has been reconstituted in lecithin liposomes and the binding of insulin investigated. A method of interpreting binding data presented as binding vs. the logarithm of free insulin concentration (binding isotherms) in terms of the binding potential concept of Wyman (1965) is described, and the results are compared with the commonly used Scatchard analysis of binding. The binding potential approach enables binding constants and Gibbs energies of formation of the insulin-receptor complex to be determined as a function of insulin bound. The limiting Gibbs energies of binding at 15 degrees C to intact cells, membranes and liposomes were found to be -55, -52 and -49 kJ mol-1 respectively. The affinity of the receptor for insulin decreases smoothly with increase in binding in all three systems. For intact adipocytes the number of insulin receptors per cell is found to be approximately 43,000.

Adipose Tissue↗

PubMed1988

Lipogenesis in rat brown adipocytes. Effects of insulin and noradrenaline, contributions from glucose and lactate as precursors and comparisons with white adipocytes.

1. Brown adipocytes were isolated from the interscapular depot of male rats maintained at approx. 21 degrees C. In some experiments parallel studies were made with white adipocytes from the epididymal depot. 2. Insulin increased and noradrenaline decreased [U-14C]glucose incorporation into fatty acids by brown adipocytes. Brown adipocytes differed from white adipocytes in that exogenous fatty acid (palmitate) substantially decreased fatty acid synthesis from glucose. Both noradrenaline and insulin increased lactate + pyruvate formation by brown adipocytes. Brown adipocytes converted a greater proportion of metabolized glucose into lactate + pyruvate and a smaller proportion into fatty acids than did white adipocytes. 3. In brown adipocytes, when fatty acid synthesis from [U-14C]glucose was decreased by noradrenaline or palmitate, incorporation of 3H2O into fatty acids was also decreased to an extent which would not support proposals for extensive recycling into fatty acid synthesis of acetyl-CoA derived from fatty acid oxidation. 4. In the absence of glucose, [U-14C]lactate was a poor substrate for lipogenesis in brown adipocytes, but its use was facilitated by glucose. When brown adipocytes were incubated with 1 mM-lactate + 5 mM-glucose, lactate-derived carbon generally provided at least 50% of the precursor for fatty acid synthesis. 5. Both insulin and noradrenaline increased [U-14C]glucose conversion into CO2 by brown adipocytes (incubated in the presence of lactate) and, in combination, stimulation of glucose oxidation by these two agents showed synergism. Rates of 14CO2 formation from glucose by brown adipocytes were relatively small compared with maximum rates of oxygen consumption by these cells, suggesting that glucose is unlikely to be a major substrate for thermogenesis. 6. Brown adipocytes from 6-week-old rats had considerably lower maximum rates of fatty acid synthesis, relative to cell DNA content, than white adipocytes. By contrast, rates of fatty acid synthesis from 3H2O in vivo were similar in the interscapular and epididymal fat depots. Expressed relative to activities of fatty acid synthase or ATP citrate lyase, however, brown adipocytes synthesized fatty acids as effectively as did white adipocytes. It is suggested that the cells most active in fatty acid synthesis in the brown adipose tissue are not recovered fully in the adipocyte fraction during cell isolation. Differences in rates of fatty acid synthesis between brown and white adipocytes were less apparent at 10 weeks of age.

Adipose Tissue↗

PubMed1999

Effect of thiazolidinediones on expression of UCP2 and adipocyte markers in human PAZ6 adipocytes.

AIMS/HYPOTHESIS: Thiazolidinediones, a new class of insulin sensitizers, up-regulate the expression of uncoupling protein 2 in rodent adipocytes. It is not known, however, whether thiazolidinediones influence uncoupling protein 2 expression in human adipocytes. We therefore investigated the effect of these drugs on uncoupling protein 2 expression in the recently immortalized human PAZ6 adipocyte cell line. METHODS: Immortalized human PAZ6 preadipocytes were differentiated into adipocytes in the presence or absence of thiazolidinediones. The effect of the drugs on uncoupling protein 2 expression and adipocyte differentiation was measured by reverse transcription-polymerase chain reaction of mRNA of uncoupling protein 2 and of five adipocyte differentiation markers. RESULTS: When cells were differentiated 15 days in the presence of thiazolidinediones, uncoupling protein 2 expression was 2.1-fold higher than in the absence of the drugs. The expression of five adipocyte differentiation markers was, however, also increased by thiazolidinediones. Short-term incubation for 4 and 24 h with thiazolidinediones increased uncoupling protein 2 expression 1.35-fold and 2.3-fold, respectively. The expression of adipocyte markers studied in parallel was also augmented. CONCLUSION/INTERPRETATION: Thiazolidinediones rapidly increase the expression of uncoupling protein 2 in human PAZ6 adipocytes but the increase of uncoupling protein 2 expression is always associated with an augmentation of the expression of all adipocyte markers studied in parallel. This indicates that the effect of thiazolidinediones on uncoupling protein 2 mRNA reflects a general increase in adipocyte differentiation rather than a specific augmentation of uncoupling protein 2 gene expression.

Adipocytes↗

PubMed2001

Human breast adipocytes express interleukin-6 (IL-6) and its receptor system: increased IL-6 production by beta-adrenergic activation and effects of IL-6 on adipocyte function.

Adipocytes produce the inflammatory cytokine interleukin-6 (IL-6); however, it is not known whether these cells express the IL-6 receptor system, how the secretion of this cytokine is regulated, and whether it has a function within adipose tissue. Using cultured human breast adipocytes, we investigated the expression of IL-6 and its receptor system, the effects of IL-6 on main adipocyte functions, and the regulation of IL-6 secretion by catecholamines and glucocorticoids. In the culture system, immunohistochemistry demonstrated expression of IL-6 and its receptor system, consisting of the ligand-binding IL-6 receptor and the signal-transducing protein gp130, in mature adipocytes, but not in undifferentiated adipocyte precursor cells. In freshly isolated adipocytes, RT-PCR detected messenger ribonucleic acids encoding the above proteins. Chronic incubation of adipocytes with 1 nmol/L IL-6 during adipose differentiation reduced glycero-3-phosphate dehydrogenase (GPDH) activity, a marker of adipocyte differentiation, and triglyceride synthesis to 67 +/- 9% of the basal level (mean +/- SEM; P < 0.05) only on day 21. Incubation of differentiated adipocytes with 10 nmol/L IL-6 for 24 h also resulted in a reduction of GPDH activity to 81 +/- 5% (P < 0.05). On the other hand, 24-h exposure to 10 nmol/L IL-6 increased basal glycerol release by 42 +/- 12% (P < 0.01) and isoproterenol-induced glycerol release by 21 +/- 6% (P < 0.05). The same concentration of IL-6, however, did not alter basal or insulin-stimulated glucose transport. IL-6 secretion was acutely and chronically stimulated by 1 micromol/L isoproterenol (peak of 6.2-fold after 3 h; P < 0.001) and only moderately suppressed by 100 nmol/L cortisol (-36 +/- 10%; P < 0.001). In conclusion, human breast adipocytes release substantial amounts of IL-6 and express IL-6 receptor and gp130. The secretion of IL-6 by adipocytes is strongly stimulated by beta-adrenergic activation and is modestly suppressed by glucocorticoids. IL-6 reduces GPDH activity and stimulates lipolysis, suggesting an autocrine/paracrine role of this cytokine in human adipose breast tissue.

Adipocytes↗

PubMed1985

Alpha 2-adrenergic binding and action in human adipocytes. Comparison between binding to plasma membrane preparations and to intact adipocytes.

Binding of the alpha 2-adrenoceptor antagonist [3H]yohimbine was demonstrated on intact human adipocytes and on human adipocyte membranes. Specific binding was rapid, reversible, saturable and of high affinity in both preparations. [3H]Yohimbine binding was inhibited by various adrenergic agents in a manner which suggests that the labeled sites probably represent the alpha 2-receptor both in intact adipocytes and in the membrane fraction. In adipocyte membranes the maximal binding capacity (Bmax) was 463 +/- 38 fmol/mg protein and the binding was of high affinity with a Kd of 2.1 +/- 0.4 nM. In intact human adipocytes, Bmax was 903 +/- 139 fmol/10(6) cells (or 342 +/- 21 fmol/100 cm2) and the binding affinity was 6.6 +/- 0.8 nM. Adrenergic antagonists bound to a homogeneous class of receptors (linear Scatchard plots) both in isolated membranes and in intact adipocytes. However, agonist binding was heterogeneous in both preparations. The affinity of agonist binding was 5-10 times higher in membranes than in intact adipocytes. The physiological relevance of the binding data was evaluated by correlating the binding of yohimbine with the antilipolytic effect of clonidine. A positive and significant correlation was found between Bmax and maximal antilipolytic effect of clonidine (r = 0.70) in membranes. Furthermore, the binding affinity (Kd) was positively correlated to the sensitivity (IC50) of the clonidine-induced antilipolysis (r = 0.65). A positive and significant correlation was also found in intact adipocytes between Bmax and the maximal antilipolytic effect of clonidine (r = 0.79). However, there was no significant correlation between Kd in intact adipocytes and the IC50 of clonidine. It is concluded that both intact human adipocytes and membrane fractions are useful models to investigate the properties and regulation of alpha 2-adrenoceptors. However, it appears from our correlation studies that the binding data obtained with the membranes are the best related to the physiological effects mediated by alpha 2-receptors in human adipocytes.

Adipose Tissue↗

PubMed2000

Ceiling culture of mature human adipocytes: use in studies of adipocyte functions.

Adipocytes contain large lipid droplets in their cytoplasm. When cultured, they float on top of the medium, clump together, and do not gain equal and sufficient access to the medium. Morphological changes cannot be observed and the majority of adipocytes undergo cell lysis within 72 h of isolation. We have used a ceiling culture method for human mature adipocytes which uses their buoyant property to allow them to adhere to a floating glass surface, where they remain viable for several weeks. Using confocal immunofluorescence microscopy we showed the cellular expression and subcellular localization of leptin in ceiling-cultured adipocytes. The secretion of leptin was increased from ceiling cultures following tumour necrosis factor-alpha treatment. Proliferation of mature human adipocytes in serum-containing medium was demonstrated by incorporation of bromodeoxyuridine, 2% of adipocytes showing positive incorporation after 4 h labelling. Proliferation was also evident from the budding of daughter cells. Apoptosis in the ceiling cultures was increased by 48 h serum deprivation (30-35 vs 10-15% in the control) and was assayed by propidium iodide staining and terminal deoxynucleotidyl transferase-mediated dUTP-fluorescein nick-end labelling. Lipolysis, analysed by liquid scintillation counting, was increased by forskolin (10 microM for 90 min) and lipogenesis, shown by autoradiography, was stimulated by insulin (10 and 100 nM for 4 h). These findings indicate that ceiling-cultured adipocytes maintain adipocyte-specific functions and that ceiling culture, which overcomes the shortcomings of adipocyte suspension culture, can be used to study adipocyte cell biology.

Adipocytes↗

PubMed1990

Expression of a 64 kD adipocyte-specific plasma membrane protein in genetically lean but not obese porcine adipocytes.

A monoclonal antibody (LA-1) to an adipocyte-specific plasma membrane protein (64 kD) was used to examine the differential expression of this protein in genetically lean and genetically obese pigs. Enzyme-linked immunosorbent assay (ELISA) implied the differential expression of the 64 kD protein in adipocyte plasma membranes having different genetic background. Sodium dodecyl-sulfate polyacrylamide gel electrophoresis (SDS-PAGE) of genetically lean, genetically obese, and contemporary subcutaneous adipocyte plasma membranes did not indicate any obvious qualitative differences in protein composition. Corresponding immunoblots utilizing LA-1 confirmed the presence of the 64 kD protein in contemporary and genetically lean adipocyte plasma membranes but absence in genetically obese adipocyte plasma membranes. LA-1 labelled intact adipocytes isolated from contemporary and genetically lean adipose tissue but did not react with isolated genetically obese adipocytes. The ability to bind to intact adipocytes indicates that the protein is exposed to the extracellular environment. The migration pattern of the protein was not affected by enzymatic deglycosylation by endoglycosidase-F suggesting that the protein is not highly, if at all, glycosylated. Presence of the 64 kD protein in genetically lean but not genetically obese adipocyte plasma membranes indicates the identification of a novel adipocyte-specific surface protein associated, either directly or secondary to the onset of obesity, with genetic predispositions for either genetically lean or obese body types in swine.

Adipose Tissue↗

PubMed1998

Stimulation of adipose differentiation related protein (ADRP) expression by ibuprofen and indomethacin in adipocyte precursors and in adipocytes.

Adipose differentiation related protein (ADRP) is a 50 kDa protein expressed at high level in differentiated adipocytes. ADRP expression is very low in undifferentiated adipocytes and increases rapidly and dramatically as the cells undergo adipose differentiation. In the present study, we demonstrate that ADRP expression at the mRNA and protein level is stimulated in adipocyte precursor cells in a time- and dose-dependent fashion by treatment with cyclooxygenase inhibitors, particularly indomethacin and ibuprofen. Lipoxygenase inhibitors such as AA861 and nordihydroguaiaretic acid were ineffective. Stimulation of ADRP expression was observed with 10(-5) M ibuprofen but maximal stimulation required a concentration of 3 x 10(-4) M. Nuclear run-on experiments indicated that indomethacin or ibuprofen stimulated the transcription of the ADRP gene in undifferentiated adipocytes. In addition to stimulating the induction of ADRP in undifferentiated cells, ibuprofen and indomethacin also stimulated the level of ADRP mRNA and protein in differentiated adipocytes. These experiments provide new information on the regulation of ADRP, an early inducible gene in the adipocyte differentiation programme in adipocyte precursors and in adipocytes and identify a new target for cyclooxygenase inhibitor action during adipocyte differentiation.

Adipocytes↗

PubMed1984

Characterization of low density lipoprotein binding to human adipocytes and adipocyte membranes.

125I-labeled low density lipoprotein (LDL) binding to purified plasma membranes prepared from freshly isolated human adipocytes was saturable, specific, and displaceable by unlabeled ligand. The maximum specific binding capacity measured at saturating concentrations of 125I-LDL was 1.95 +/- 1.17 micrograms of LDL bound/mg of membrane protein (mean +/- S.D., n = 16). In contrast to cultured fibroblasts, specific binding of LDL to adipocyte membranes was calcium-independent, was not affected by EDTA or NaCl, and was not destroyed by pronase. Plasma membranes purified directly from homogenized adipose tissue also showed calcium-independent LDL specific binding (0.58 +/- 0.33 micrograms of LDL bound/mg of membrane protein, mean +/- S.D. n = 11). Specific binding, internalization, and degradation of 125I-methylated LDL was demonstrated in isolated adipocytes and competition experiments showed that native and methylated LDL interacted with adipocytes through some common recognition mechanism(s). Compared to native LDL, specific binding of methylated LDL to adipocyte membranes was significantly reduced (43%), indicating that interaction of LDL with adipocyte was dependent in part on the lysine residues of apolipoprotein B. LDL binding to adipocyte plasma membranes was also competitively inhibited by human high density lipoprotein subfractions HDL2 and HDL3. Thus, LDL metabolism in mature adipocytes appears to be regulated by mechanisms distinctly different from a variety of cultured mesenchymal cells. In addition, the ability of adipocytes to bind, internalize, and degrade significant amounts of methylated LDL supports the view that adipose tissue is involved in the metabolism of modified lipoproteins in vivo.

Adipose Tissue↗

PubMed1981

Response of white adipocyte of mouse and rabbit to catecholamines and ACTH. 2. Stability and restoration of activity of hormone-sensitive adenylate cyclase of adipocyte plasma membrane.

The isolated intact white adipocyte of the Swiss mouse responds to both ACTH and catecholamines by an elevation of cAMP levels and an increase in lipolysis. However, in the isolated plasma membrane of the mouse adipocyte, adenylate cyclase loses its responsiveness to ACTH but retains its ability to respond to catecholamines. This lack of responsiveness to ACTH by adenylate cyclase of mouse adipocyte plasma membrane can be overcome, at least partially, by addition of GPP (NH)p, an analog of GTP, to the assay medium. The data on mouse adipocyte membrane suggests that the coupling of ACTH receptor to adenylate cyclase is dependent on GTP and that catecholamine-activation of adenylate cyclase is less dependent on this nucleotide. The isolated intact white adipocyte of adult New Zealand rabbit responds to ACTH, but does not (or only weakly) respond to catecholamines. In contrast to the mouse plasma membrane preparation, adenylate cyclase of adipocyte membrane of the rabbit responds to ACTH. And the addition of GPP(NH)P is not required to demonstrate the CTH: sensitive adenylate cyclase activity. The difference between mouse and rabbit adipocyte membrane in the requirement for GPP(NH)P in ACTH action is not readily explained. The lack of catecholamine sensitivity of rabbit membrane enzyme cannot be reversed by addition of GPP(NH)P or adenosine deaminase. These two adenylate cyclase model systems using mouse and rabbit adipocyte plasma membrane may be useful tools for the study of the specificity and mechanism of action of lipolytic hormones such as ACTH and catecholamines.

Adenylyl Cyclases↗

PubMed1988

Characterization of high-density lipoprotein binding to rat adipocytes and adipocyte plasma membranes.

The interaction of high-density lipoproteins (HDL) with adipocytes is important in the regulation of cellular cholesterol flux. To study the mechanisms of HDL binding and cellular processing, we incubated adipocytes isolated from epididymal and perirenal adipose tissue of male Wistar rats (300 g) with HDL1 (1.07-1.10 g/mL) and HDL2 (1.10-1.14 g/mL) fractions separated from rat plasma by gradient ultracentrifugation. Freshly isolated adipocytes were incubated with 125I-labeled HDL for 2 h at 37 degrees C to determine cell-associated uptake and degradation. Adipocytes from both fat regions showed significant cell-associated HDL1 and HDL2 uptake and very high medium degradation (2- to 6-fold higher than uptake). To assess 125I-labeled HDL binding independent of cellular metabolism, we purified adipocyte plasma membranes from isolated adipocytes and used them in binding assays. Binding of HDL1 and HDL2 in the membrane system was 85-95% specific, sensitive to high NaCl concentrations, and abolished by pronase treatment. In contrast to HDL2 binding, the maximum HDL1 binding to perirenal plasma membranes was significantly higher than its binding to epididymal membranes (7.2 +/- 1.3 vs. 4.4 +/- 0.2 micrograms/mg, n = 6, p less than 0.05). This increment in HDL1 binding to perirenal membranes represented an EDTA- sensitive, calcium-dependent component. These results indicate that HDL binding to adipocyte plasma membranes depends on both adipose tissue region and HDL subtype. The membrane binding characteristics, taken together with the cellular uptake results, suggest that adipocytes bind and metabolize HDL and that this interaction may involve a protein receptor.

Adipose Tissue↗

PubMed1985

[Studies on adipocyte glucagon receptor assay--with special reference to the effect of cold acclimation on glucagon receptors of white adipocytes].

The changes in glucagon receptors of white adipocytes from cold-acclimated rats were investigated to know the metabolic role of glucagon in cold acclimation by establishing a glucagon radioreceptor assay system for isolated white adipocytes. Glucagon radioreceptor assay methodology The binding of 125I-labelled glucagon to isolated epididymal white adipocytes was linearly related to the number of cells (0.5-2.0 X 10(5) cells/ml) added in the medium. At a cell concentration higher than 3.0 X 10(5) cells/ml, the amount of specific binding failed to show the proportional relationship to the number of adipocytes. The effects of incubation temperature (4 degrees C, 25 degrees C and 37 degrees C) on the glucagon binding were investigated. Incubation at 25 degrees C was adopted in the present study because of the highest maximum binding and the longest steady state obtained. Preincubation at 25 degrees C for 15 min increased significantly the amount of specific binding. It was confirmed that bacitracin, polypeptide antibiotics, inhibited significantly the degradation of glucagon. The glucagon binding under these conditions was found to be saturable and reversible, validating a specific reaction for the glucagon receptor. When a Scatchard plot was constructed, the data was curvilinear with an upward concavity, indicating the presence of at least two classes of binding site with different fixed affinities or of negatively cooperative interactions between receptors. It was concluded that an appropriate condition for glucagon receptor assay of white adipocytes consists of cell concentration of 1 X 10(5) cells/ml, 15 minute-preincubation and 30 minute-reaction at 25 degrees C in the presence of bacitracin (1 mg/ml). Effect of cold acclimation on glucagon receptors of white adipocytes Cold acclimation decreased the size and increased the number of epididymal white adipocytes. Cold acclimation increased the number of glucagon receptors of white adipocytes; about 140% increase expressed as per cell, approximately 260% increase per unit of surface area and 210% increase per whole tissue. The affinity of binding sites was not changed. The increased binding sites could explain, at least partly, the enhanced metabolic response of cold-acclimated rats to glucagon.

Acclimatization↗

PubMed1983

A trypsin-sensitive, heat-labile, N-ethylmaleimide-sensitive factor in adipocyte post-microsomal supernatant which affects the assay of adipocyte glycerol phosphate acyltransferase activities.

Addition of adipocyte 100 000 g post-microsomal supernatant to assays of glycerol phosphate acyltransferase in isolated mitochondria or microsomal fractions decreased activity at lower concentrations of palmitoyl-CoA. At higher concentrations of palmitoyl-CoA, activation was observed on addition of post-microsomal supernatant. The effect of post-microsomal supernatant to decrease activity at lower [palmitoyl-CoA] was abolished by heating or by trypsin treatment, and was also abolished by addition of N-ethylmaleimide to assays or by pretreatment of post-microsomal supernatant with N-ethylmaleimide. The stimulatory effect seen at higher [palmitoyl-CoA] was not sensitive to heat or trypsin treatment. The effect of post-microsomal supernatant at lower [palmitoyl-CoA] cannot be attributed to palmitoyl-CoA hydrolase activity. It was found that brief treatment of adipocyte mitochondria with low concentrations of trypsin was an effective way to remove contaminating microsomal glycerol phosphate acyltransferase activity. Adipocyte post-microsomal supernatant was more effective than an equivalent quantity of liver post-microsomal supernatant protein in decreasing adipocyte microsomal glycerol phosphate acyltransferase activity. The effects of the supernatants from both tissues were decreased by flavaspidic acid. Semi-purified Z-protein fraction from rat liver did not mimic the effect of adipocyte post-microsomal supernatant to decrease glycerol phosphate acyltransferase at lower [palmitoyl-CoA]. Post-microsomal supernatants obtained from noradrenaline-treated adipocytes were less effective than those from control cells in decreasing glycerol phosphate acyltransferase activity in microsomal fractions at lower [palmitoyl-CoA]. It is suggested that adipocyte cytosol may contain an acyl-CoA-binding protein or proteins differing from Z-protein in some respects. The physiological significance of the findings is briefly discussed.

Acyltransferases↗

PubMed1986

Conversion to adipocytes of a clonal bone marrow preadipocyte line (H-1/A) and fatty acid composition of the resultant adipocytes.

Conversion to adipocytes and fatty acid composition were investigated in a clonal bone marrow preadipocyte line (H-1/A). The growing cells exhibited a fibroblastic appearance. After the cessation of growth, triacylglyceride (TG) synthesis in the cells increased as they incorporated precursor from the growth medium and became adipocytes. Hydrocortisone and insulin accelerated the TG synthesis in H-1/A cells in a dose-dependent manner when they were cultured in the growth medium containing 10% horse serum. The rate of conversion to adipocytes was reduced as the concentration of horse serum was decreased, and this reduction was not influenced by the addition of insulin and/or hydrocortisone. These results suggest that conversion to adipocytes of H-1/A cells is primarily dependent on some component(s) of the serum. Conversion to adipocytes of the cells may involve a process of differentiation since the conversion was completely inhibited when the cells were cultured in the presence of bromodeoxyuridine. Fatty acid composition was significantly different between adipose H-1/A cells and adipocytes derived from other marrow preadipocyte line MC3T3-G2/PA6 cells. Unsaturated fatty acids accounted for 76% of the fatty acid composition of adipose H-1/A cells; in contrast, saturated fatty acids constituted 65% of the fatty acid composition of the adipose MC3T3-G2/PA6 cells. These results suggest that there is a heterogeneity of preadipocytes in bone marrow. These two preadipocyte lines thus provide a useful tool for the study of marrow adipocytes and can also be used to analyze the hematopoietic microenvironment through studies of the effect of these cells on hematopoietic cell proliferation.

Adipose Tissue↗

PubMed1986

Studies on the expression of adipocyte-specific cell surface antigens during the differentiation of adipocyte precursor cells in vitro.

Using species and cell specific antiadipocyte sera an immunoprecipitation procedure was developed which allowed the nature of adipocyte cell surface antigens to be investigated. Analysis of immunoprecipitates from mature adipocyte plasma membranes of rat, ox and chicken and similar 125I-labelled membranes revealed the presence of specific externally disposed adipocyte specific antigens which were also species specific. For mature cells the specific antigens had molecular weights of 124,000, 92,000 and 59,000 in the case of the rat, 87,000 in the case of the ox and 56,000, 47,000 and 37,000 in the case of the chicken. None of these antigens were cross immunoprecipated by antisera to non-homologous adipocytes. The presence of the antigens at the surface of differentiating rat while adipocyte precursor cells was demonstrated using a labelled-second antibody cellular immunoassay and the expression of this reactivity revealed to be an early event in the differentiation programme of the cells. The increase in cell surface immunoreactivity during the differentiation of the cells was shown to be dependent upon the expression of two of the antigens previously shown to be markers of the mature adipocyte phenotype. The functional identity and possible role of these antigens in the control of adipocyte differentiation in vitro and in vivo now becomes accessible to investigation experimentally.

Adipose Tissue↗

PubMed1995

Functional assessment of white and brown adipocyte development and energy metabolism in cell culture. Dissociation of terminal differentiation and thermogenesis in brown adipocytes.

We investigated the effect of insulin, triiodothyronine (T3) and dexamethasone (a synthetic glucocorticoid) on differentiation, lipid metabolism and thermogenesis of preadipocytes isolated from white fat (WAT) and brown fat (BAT) from the Siberian dwarf hamster (Phodopus sungorus). Cell cultures from WAT and BAT were chronically treated with the above hormones alone or in any combination. After differentiation (day 8 or 9 of culture) we measured the following parameters: adipogenic index (number x size of adipocytes), protein content, lipolysis, cell respiration, and expression of the uncoupling protein UCP, which is unique to mitochondria of brown adipocytes. Insulin was the most important adipogenic factor for brown and white adipocytes and necessary for terminal differentiation, whereas dexamethasone alone completely inhibited differentiation. T3 had no effect on adipogenesis in WAT cultures, but further increased insulin stimulated adipogenesis in BAT cultures. Basal lipolysis was higher in WAT than in BAT cultures except when dexamethasone was present, which stimulated lipolysis in both culture types to the same extent. T3 had a pronounced dose dependent lipolytic effect on WAT cultures but very little effect on BAT cultures. Respiration rates were generally higher in differentiated adipocytes than in fibroblast like cells. T3 had no effect on thermogenesis in WAT cultures but increased thermogenesis in BAT cultures, and this was further elevated by insulin. UCP expression in BAT cultures could be detected by western blot in insulin treated, T3 treated and insulin+T3 treated cultures with highest expression in the latter. These results imply a possible dissociation of terminal differentiation and thermogenic function of brown adipocytes. In WAT cultures there was also a low level of UCP detectable in the insulin+T3 treated cultures. Immuno-fluorescence microscopy analysis revealed the presence of UCP in 10-15% of adipocytes from WAT cultures (in BAT cultures: 90%), indicating the presence of some brown preadipocytes in typical WAT deposits.

Adipocytes↗

PubMed2000

Adipocyte metabolism in adipocyte fatty acid binding protein knockout mice (aP2-/-) after short-term high-fat feeding: functional compensation by the keratinocyte [correction of keritinocyte] fatty acid binding protein.

Mice null for adipocyte fatty acid binding protein (AFABP) compensate by increasing expression of keratinocyte fatty acid binding protein (KFABP) (Hotamisligil et al. Science 274:1377-1379, 1996). In the present study, AFABP knockout (KO) and wild-type (WT) mice became equally obese on a high-fat diet, as judged by fat pad weights, adipocyte size, and body composition analysis. High-fat feeding led to moderate insulin resistance in both WT and AFABP knockout mice, as indicated by an approximately 2-fold increase in plasma insulin. However, in the high fat-fed mice, plasma glucose levels were approximately 15% lower in the AFABP-KO mice. Adipocytes isolated from AFABP-KO and WT mice fed high- or low-fat diets exhibited similar rates of basal and norepinephrine-stimulated lipolysis and insulin-stimulated rates of glucose conversion to fatty acids and glyceride-glycerol. However, basal glucose conversion to fatty acids was higher in adipocytes of AFABP-KO mice. Adipocyte tumor necrosis factor-alpha release was similarly increased by high-fat diet-induced obesity in both WT and AFABP-KO mice. As assessed by Western blot analysis, the level of KFABP protein in AFABP-KOs was approximately 40% of the level of AFABP in WT controls. The binding affinities of KFABP for long-chain fatty acids were 2- to 4-fold higher than those of AFABP, but the relative affinities for different fatty acids were similar. As for AFABP, the rate of fatty acid transfer from KFABP to model phospholipid vesicles was increased with acceptor membrane concentration and by inclusion of acidic phospholipids, indicating a similar mechanism of transfer. We conclude KFABP can functionally compensate for the absence of AFABP, resulting in no major alterations in adipocyte metabolism or fat accumulation in response to short-term feeding of high-fat diets that result in moderate hyperinsulinemia.

Adaptation, Physiological↗