Growth of preadipocyte cell lines and cell strains from rodents in serum-free hormone-supplemented medium

Summary Ob17 is a clonal cell line isolated from the epididymal fat pad of C57 BL/6J ob/ob mouse that differentiates into adiposelike cells in serum-supplemented medium. In serum-free medium, this cell line shows increased growth under the addition of insulin, transferrin, fibroblast growth factor (FGF), and a factor present in extract of rat submaxillary gland (SMGE). This medium is referred to as 4F. Epidermal growth factor or nerve growth factor cannot replace SMGE, whereas partially purified platelet extract can substitute for FGF but only partially for SMGE. 4F Medium is able to support the proliferation of cells from other established preadipocyte clonal lines, HGFu and 3T3-F442A, and also of preadipocyte cells isolated from the stromal-vascular fraction of rat and mouse adipose tissues. In each case 4F medium is insufficient to support the differentiation of these cells into adipocytes. Ob17 cells grown and maintained in serum-free hormone-supplemented medium retain the ability to convert to adiposelike cells after serum addition. This serum requirement for differentiation cannot be substituted by the addition of growth hormone or of other putative adipogenic factors, or both. The results are discussed with respect to the requirements for growth and differentiation of the 3T3-L1 and 1246 preadipocyte cell lines previously described. This work was supported by the “Centre National de la Recherche Scientifique” (Grant 1208-Biochimie du Développement and Grant 4162-Endocrinologie), by the “Ministère de la Recherce et de la Technologie” (Grant 81-L-1322), by the “Fondation pour la Recherche Médicale,” by NATO (Grant 1704), and by the “Institut National de la Santé et de la Recherche Médicale” (Grant 827006).     

Transformation of Ob17 cells promotes proliferation and differentiation of Ob17 preadipocytes via distinct extracellular intermediates

Conditioned serum-free medium of Ob17 cells transformed by the middle-T-only gene of polyoma virus (Ob17MT cells) is able to support growth and adipose conversion of the parental Ob17 cells. Conditioned media from 3T3-F442A cells (untransformed preadipocyte clonal line) and MTT4 cells (middle-T-transformed non-preadipocyte clonal line) are inactive. The serum-free conditioned medium of Ob17MT cells is also active on growth and adipose conversion of 3T3-F442A cells. The morphological differentiation of Ob17 cells is accompanied by the expression of early (lipoprotein lipase, LPL) and late (glycerol-3-phosphate dehydrogenase, GPDH) biochemical markers of adipose conversion. Bio-Gel P-60 chromatography and SDS-PAGE have allowed characterization of a mitogenic fraction of apparent MW approximately equal to 28 Kd distinct from an adipogenic fraction of apparent MW less than 10 Kd. This adipogenic fraction is only required for the acquisition of the GPDH activity and is therefore active on terminal differentiation.     

Isolation of myoblastic, fibro-adipogenic, and fibroblastic clonal cell lines from a common precursor and study of their requirements for growth and differentiation

C17-S1-D-T984 (to be referred to as T984) is a myogenic clonal cell line isolated from a mouse teratocarcinoma. T984 exhibits phenotypic instability since it gives rise not only to myogenic but also to fibro-adipogenic and fibroblastic clones. A cell line of each clone type has been established and studied with respect to (1) phenotypic expression and stability; and (2) growth and differentiation in serum-free and serum-supplemented media. In both respects, marked differences between the three cell lines were observed. All three cell lines respond by increased growth in serum-free media to insulin, transferrin, fibroblast growth factor (FGF) and the serum-spreading factor of Holmes. The fibroblastic and the fibro-adipogenic cell lines can both be grown indefinitely in a serum-free medium which contains the above factors. The fibro-adipogenic cell line, which differentiates in serum-supplemented medium, exhibits very limited differentiation in the absence of serum; the serum factor(s) required for adipogenic differentiation is (are) probably proteins of molecular weight superior to 10 000. In direct contrast, the myogenic cell line exhibits limited growth in serum-free medium but readily differentiates under these conditions. Moreover, myogenic differentiation could be obtained in the defined medium at very low densities and was not influenced by the addition of medium conditioned by cells seeded at high densities. Thus, in this system, muscular differentiation is apparently independent of diffusible endogenous or exogenous factors and is probably triggered by the arrest of growth. While our results do not explain the reason why T984 exhibits phenotypic instability, they do indicate that this clonal cell line and its clonal derivatives could be used to identify the factors that influence the growth and the differentiation of cells of different mesenchymal phenotypes. The possible relationship of phenotypic instability to muscular dystrophies is also discussed.     

Protein factor obtained from rat adipose tissue specifically permits the proliferation of the 3T3-L1 and Ob1771 cell lines

We have found the presence of protein factor in rat adipose tissue which permits the proliferation of 3T3-L1 and Ob1771 preadipocytes cultured in a completely defined serum-free medium containing only progression factors [epidermal growth factor (EGF) and insulin] as growth factors. This mitogenic activity of the protein factor was not detected in various other cell lines, in particular, Swiss 3T3 cells which could proliferate in response to a competent factor [platelet-derived growth factor (PDGF) or fibroblast growth factor (FGF)] in the same serum-free medium. This activity of the factor was heat- and pronase-unstable, and reductant-stable, and the apparent molecular weight of the factor was about 20,000. These results strongly suggest that the protein factor is different from PDGF or FGF and contributes to the formation of new adipocytes by specifically stimulating the proliferation of preadipocytes, acting like competent factor.     

Induction of oligodendrocyte-like properties in a primitive hypothalamic cell line by cholesterol, an eye derived growth factor and brain extract

A serum-free medium has been devised which permits proliferation of the mouse primitive nervous cell line F7. When cholesterol, eye-derived growth factor and brain extract are added in this medium for 48 h, 80-90% of oligodendrocyte-like cells are generated. These cells have diminished substrate adhesion. They acquire the capacity to synthesize carbonic anhydrase II and myelin basic protein, two specific proteins of oligodendrocytes. These observations suggest that F7 clonal cell line, which has been previously shown to be a neurophysin cell precursor, is also a precursor for oligodendrocytes, and represents a bipotent stem cell line for both neuronal and glial cell lineages.     

Defined medium for normal adult human prostatic stromal cells

Summary Stromal-epithelial interactions are pivotal in many aspects of prostatic biology. A defined culture system is critical for the investigation of factors that regulate the growth and differentiation of human prostatic stromal cells. We have identified conditions which promote stromal cell attachment and proliferation in serum-free medium. MCDB 201, originally developed for the clonal growth of chick embryo fibroblasts, proved to be a superior basal medium of those that we tested. Supplementation of MCDB 201 with basic fibroblast growth factor (FGF), insulin-like growth factor (IGF), and platelet-derived growth factor (PDGF) permitted attachment and exponential growth of cells throughout a 7-d period with an initial inoculum as low as 10³ cells per well of a 96-well microtiter dish. Using these assay conditions, we subsequently verified that basic FGF and IGF, but not PDGF, were required for optimal growth. No activity was found for heparin, transferrin, or the androgen R1881. Epidermal growth factor (EGF) didn’t stimulate growth when added to medium containing basic FGF and IGF, but was moderately stimulatory when added to basal medium alone. Cholera toxin inhibited growth. This simple and efficient culture medium provides a suitable assay system for more extensive studies of growth regulation and differentiation of human prostatic stromal cells, and will provide the basis for future development of a defined medium that supports clonal growth. Characterization of stromal-epithelial interactions will be facilitated by the use of this defined culture system for stromal cells in conjunction with the serum-free culture systems previously developed for human prostatic epithelial cells.     

Improvement in the proliferative activity of human-human hybridomas at low cell density by transfection with bFGF gene

Highly purified recombiaant basic fibroblast growth factor (rbFGF) and acidic FGF (aFGF) stimulated the proliferation of human-human (h-h) hybridomas to the extent of over four-fold from a low cell density such as 1×10³ cells per ml in a serum-free medium in 24-well plates. The stimulatory effect of rbFGF was also observed in various lymphoid cell lines. Expecting that FGF could be an autocrine growth factor, we introduced bFGF gene into a h-h hybridoma using an expression plasmid induced by dexamethasone. The transformed cells thus obtained, HPO-75.11bbFGF-7, were able to grow well from a low inoculum density in a serum-free medium and antibody production was also increased when bFGF gene expression was induced. The transformed cells could grow at clonal density in a serum-free medium in 96-well plates, though the original cells could not. We also obtained a more practical transfectant, HPO-75.29-H74, using a high-shear stress adapted clone as the recipient and an expression plasmid having bFGF gene under the control of metallothioneine-I promoter. The HOP-75.29-H74 cells were capable of growing and producing human monoclonal antibody against hepatitis B virus surface antigen from an inoculum density of 1×10³ cells per ml in an agitation vessel without addition of an inducer.     

Relationship between androgen-induced cell proliferation and sensitivity to exogenous growth factors

The relationship between growth factor responses and androgen-induced cell proliferation was studied in a mouse renal tumor (RAG) cell line, a hybrid (F614B16) rat prostate x RAG cell line, and an 8-azaguanine-resistant revertant of the F614B16 cell line. The hybrid F614B16 cells are very sensitive to androgens; treatment with 20 nM 5 alpha-dihydrotestosterone accelerated cell growth in the presence or absence of serum. In contrast, the RAG cells and 8-azaguanine-resistant F614B16 cells responded to 5 alpha-dihydrotestosterone only in the absence of serum. Variation in the proliferative response to androgens among these cell lines was associated with variation in growth factor sensitivity. Basic fibroblast growth factor (bFGF) stimulated basal and androgen-induced growth of F614B16 cells in serum-free and serum-supplemented media, whereas it inhibited RAG cell growth. Basic FGF stimulated basal, but not androgen-induced growth of revertant F614B16 cells. The cell lines also differed in sensitivity to epidermal growth factor, which had no effect on hybrid cell growth but inhibited RAG and revertant cell growth in a dose-dependent fashion in serum-free media. The results of these studies suggest that androgen-sensitivity is associated with a positive response to FGF and insensitivity to exogenous epidermal growth factor.     

Growth-stimulatory effects of androgen, high concentration of glucocorticoid or fibroblast growth factors on a cloned cell line from Shionogi carcinoma 115 cells in a serum-free medium

The effects of various kinds of growth factors or steroids on the proliferation of Shionogi carcinoma 115 (SC115) cells were investigated in cell culture. In a serum-free medium [Ham's F-12:Eagle's minimum essential medium (1:1, vol/vol) containing 0.1% bovine serum albumin], the proliferation of SC-3 cells (a cloned cell line from SC115 cells) estimated by [3H]thymidine incorporation into DNA and cell number reached a plateau at 10(-8) M testosterone (up to 200-fold), 10(-7) M dexamethasone (up to 30-fold) or 1 ng/ml of fibroblast growth factors (FGF; up to 50-fold). However, the proliferation in the serum-free medium was not significantly stimulated by the addition of low to very high concentrations of progesterone, oestradiol-17 beta, epidermal growth factor, platelet derived growth factor or insulin; transforming growth factor beta slightly stimulated the growth (up to 5-fold) but markedly inhibited the growth stimulation induced by testosterone. Furthermore, an epithelial appearance of SC-3 cells grown in the absence of growth factors or steroids was changed to a fibroblast-like appearance only by the addition of testosterone, high concentrations of dexamethasone or FGF. By investigating various kinds of growth factors or steroids, the present study demonstrates that androgen, high concentration of glucocorticoid or FGF alone significantly stimulates the proliferation of SC-3 cells with a change of morphology in the serum-free medium.     

Growth of myoblasts in lipoprotein-supplemented,serum-free medium: Regulation of proliferation by acidic and basic fibroblast growth factor

Summary BC₃H1 myoblast cells seeded at low density on gelatin-coated dishes and exposed to a 1∶1 (vol/vol) mixture of Dulbecco’s modified Eagle’s medium and Ham’s F12 medium, proliferate actively when exposed to high density lipoproteins (HDL), transferrin, insulin, and basic or acidic fibroblast growth factor (FGF). This serum-free medium combination supported cell multiplication at a rate equal to that of serum-supplemented medium, and at low cell input (10³ cells/35-mm dish). It also allowed serial transfer of the cultures under serum-free conditions. HDL seems to promote cell survival and to act as progression factor allowing cells to divide when exposed to either basic or acidic FGF. When the potency of basic and acidic FGF were compared, acidic FGF was 20-fold less potent than basic FGF.     

Regulation of rat ovarian cell growth and steroid secretion

A cultured rat ovarian cell line (31 A-F(2)) was used to study the effect of growth factors (epidermal growth factor [EGF] and fibroblast growth factor [FGF]), a survival factor (ovarian growth factor [OGF]), a hormone (insulin), and an iron-binding protein (transferring) on cell proliferation and steroid production under defined culture conditions. EGF and insulin were shown to be mitogenic (half-maximal response at 0.12 nM and 0.11 muM, respectively) for 31A-F(2) cells incubated in serum-free medium. EGF induced up to three doublings in the cell population, whereas insulin induced an average of one cell population doubling. FGF, OGF, and transferrin were found not to have any prominent effect on cell division when incubated individually with 31A-F(2) cells in serum-free medium. However, a combination of EGF, OGF, insulin, and transferrin stimulated cell division to the same approximate extent as cells incubated in the presence of 5 percent fetal calf serum. EGF or insulin did not significantly affect total cell cholesterol levels (relative to cells incubated in serum-free medium) when incubated individually with 31A-F(2) cells. However, cell cholesterol levels were increased by the addition of OGF (250 percent), FGF (370 percent), or a combination of insulin and EGF (320 percent). Progesterone secretion from 31A-F(2) cells was enhanced by EGF (25 percent), FGF (80 percent), and insulin (115 percent). However, the addition of a mitogenic mixture of EGF, OGF, insulin, and transferrin suppressed progesterone secretion 150 percent) below that of control cultures. These studies have permitted us to determine that EGF and insulin are mitogenic factors that are required for the growth of 31A-F(2) cells and that OGF and transferrin are positive cofactors that enhance growth. Also, additional data suggest that cholesterol and progesterone production in 31A-F(2) cells can be regulated by peptide growth factors and the hormone insulin.     

Adipose cell differentiation in culture

Summary The isolation of preadipocyte cell strains from adipose tissue and from bone marrow, and the establishment of preadipocyte cell lines from embryonic and adult mouse, have been useful tools to study the process of adipose cell differentiation.This process is regulated both by extracellular signals present in serum and by intracellular signals; the characterization of these signals is under investigation. During adipose cell differentiation morphological and enzymatic changes are dramatic and they are accompanied by qualitative and quantitative variations of the cell protein content. These changes include the induction of the enzymes of the fatty acid and triglyceride synthesizing pathways and a subsequent triglyceride accumulation. The development of hormonal responses to insulin and to -adrenergics is also observed, and differentiated adipose cells behave essentially like mature adipocytes isolated from adipose tissue.The present review will be devoted to the main events of adipose conversion in cell lines and cell strains, and to current work which concerns the identification of the triggering signals possibly involved in that process.Abbreviations LPL lipoprotein lipase - MGL monoglyceride lipase - T3 triiodothyronine - dbcAMP dibutyryl-CAMP - MIX 1-methyl-3-isobutylxanthine - PG prostaglandins - FCS fetal calf serum - EDGF eye-derived growth factor - PDGF platelet-derived growth factor - FGF fibroblast growth factor - VLDL very low density proteins     

Neoplastic progression of rat tracheal epithelial cells is associated with a reduction in the number of growth factors required for clonal proliferation in culture

Summary Factors regulating the proliferation of normal, preneoplastic, and neoplastic rat tracheal epithelial (RTE) cells were investigated to identify changes taking place during the progression of RTE cells to neoplasia. Normal RTE cells exhibit clonal proliferation in a serum-free medium containing pituitary extract, serum albumin, cholera toxin, epidermal growth factor, hydrocortisone, and insulin. All combinations of these six factors were examined for their abilities to support clonal proliferation of normal, preneoplastic, and neoplastic RTE cells. In general, preneoplastic RTE cells required fewer factors for proliferation than normal RTE cells, and neoplastic cells required fewer factors than preneoplastic cells. A common pattern of reductions has been identified in the growth factors required for the clonal proliferation of preneoplastic vs. normal RTE cells and for neoplastic vs. preneoplastic and normal RTE cells. Normal RTE cells exhibit clonal proliferation in a serum-free medium supplemented with a minimum of six factors: bovine serum albumin, bovine pituitary extract, cholera toxin, epidermal growth factor, hydrocortisone, and insulin. Preneoplastic RTE cells exhibit clonal proliferation in a serum-free medium supplemented with four factors: bovine serum albumin, bovine pituitary extract, hydrocortisone, and insulin. Finally, neoplastic RTE cells exhibit clonal proliferation in a serum-free medium supplemented with two factors: bovine serum albumin and bovine pituitary extract. These results suggest that the progression of RTE cells to neoplasia is associated with a series of changes in regulatory pathways that control cell proliferation.     

Partial expression of monoaminergic (serotoninergic) properties by the multipotent hypothalamic cell line F7. An example of learning at the cellular level

A serum-free medium supplemented with a glial conditioned medium, a brain extract from 8-to 10-day-old mice, hormones, and eye-derived growth factor has been devised which permitted the mouse primitive hypothalamic nerve cell line F7 to express some biochemical properties typical of monoaminergic neurons. Maximal expression was obtained when the culture conditions were applied for 2 days. Most (90–95%) cells then synthesized [³H]serotonin from [⁴H]5-hydroxytryptophan (but not from [³H]tryptophan). No synthesis was detected in the presence of carbidopa (20 μM), therefore suggesting the involvement of l-aromatic-amino-acid decarboxylase in this process. In addition, F7 cells cultured in such serum-free medium exhibited the capacity of accumulating exogenous serotonin by an ouabain-sensitive mechanism. These data further supported that active molecules in the cell environment can induce, in a primitive cell line, some of the enzymatic activities associated with monoaminergic neurons. Since other well-defined culture conditions can promote the differentiation of the same clone into oligodendrocytes (De Vitry et al., 1983), it can be concluded that the F7 cell has the properties of an embryonic stem cell of the CNS which, depending on external signals, may switch into different alternative developmental neural pathways. We postulate that the stabilization of neuron-like properties due to repetitive cell stimulation by active signals in the environment may represent an example of learning at the cellular level.     

Mouse epidermal keratinocytes. Clonal proliferation and response to hormones and growth factors in serum-free medium

A serum-free medium (LEP-1) has been developed for mouse epidermal keratinocytes. LEP-1 consists of "Ca2+-free" Eagle's MEM with non-essential amino acids and seven added supplements (transferrin, 5 micrograms/ml; epidermal growth factor (EGF), 5 ng/ml; hydrocortisone, 0.5 microM; insulin, 5 micrograms/ml; phosphoethanolamine and ethanolamine, each 50 microM; bovine pituitary extract, 180 micrograms of protein/ml). Although serum-free the culture system was dependent for growth on bovine pituitary extract as the only still undefined supplement. LEP-1 supports sustained multiplication of mouse keratinocytes for 25 or more population doublings. A clonal growth assay was developed to investigate the action of growth factors, hormones and other supplements on keratinocytes. Cells grown in LEP-1 (calcium concentration was 0.03 mM) maintained a high proliferative rate and presented the typical morphology of basal epidermal cells. When the calcium concentration of the medium was raised to 1.0 mM, the cells were triggered to differentiate terminally. The epithelial nature of the cells was demonstrated both by electron microscopy and by immunostaining with anti-keratin antibody. The maturation stage of the keratinocytes was defined by several morphological features during the proliferative phase and in terminally differentiating cultures. This serum-free system supported a useful number of cell divisions while keratinocytes retained the capacity to undergo terminal differentiation when given the appropriate stimulus. It provides, therefore, provides a useful model for investigations on growth, differentiation and malignant transformation of epidermal cells in culture.     

Roles of various growth factors in growth of human osteosarcoma cells which can grow in protein-free medium.

The human osteosarcoma cell line (OST-1-PF) can grow in protein-free Coon's modified Ham's F12 medium. Growth of the cells in protein-free medium was partially density-dependent and partially depressed by medium change. An extract and conditioned medium of OST-1-PF cells contained high mitogenic activity for BALB/c3T3 cells. The growth factor in the cells was purified and identified as a basic fibroblast growth factor (bFGF)--like factor on the basis of its elution profile on heparin-affinity chromatography and the result of immunoblotting. An unidentified factor in a conditioned medium eliciting most of the DNA synthesis-stimulating activity showed a weak affinity for heparin. Various additions, including serum and growth factors, stimulated the growth of OST-1-PF cells in protein-free medium. Of these factors, epidermal growth factor (EGF), transforming growth factor-alpha (TGF-alpha), acidic fibroblast growth factor (aFGF) and bFGF were the most potent mitogens. High-affinity receptors of EGF and FGF were found on the surface of these cells. These results indicate that autonomous growth of OST-1-PF cells in protein-free medium is mainly controlled by an intracellular mechanism.     

Control by fibroblast growth factor of differentiation in the BC3H1 muscle cell line

The regulation of creatine phosphokinase (CPK) expression by polypeptide growth factors has been examined in the clonal mouse muscle BC3H1 cell line. After arrest of cell growth by exposure to low concentrations of serum, BC3H1 cells accumulate high levels of muscle-specific proteins including CPK. The induction of this enzyme is reversible in the presence of high concentrations of fetal calf serum, which cause quiescent, differentiated cells to reenter the cell cycle. Under these conditions, the rate of CPK synthesis is drastically reduced. We show in the present communication that either pituitary-derived fibroblast growth factor (FGF) or brain-derived FGF are as effective as serum in repressing the synthesis of CPK when added to quiescent, differentiated cells. The decrease in the rate of synthesis of CPK occurs within 22 h after the addition of pituitary FGF to the cells. Pituitary FGF had very little effect, if any, on the rate CPK degradation. The overall rate of protein synthesis and the pattern of synthesis of the major polypeptides made by these cells was not altered by the addition of FGF. Although pituitary FGF was mitogenic for BC3H1 cells, the rate of cell growth was not absolutely correlated with the extent of repression of CPK. Brain-derived FGF fully repressed CPK induction under conditions where it showed no significant mitogenic activity. These results show that the expression of a muscle-specific protein, CPK, can be controlled by a single defined polypeptide growth factor in fully differentiated cultures, and that initiation of cell division is not required for their regulation to take place.     

Regulation of skeletal muscle satellite cell proliferation and differentiation by transforming growth factor-beta, insulin-like growth factor I, and fibroblast growth factor

Skeletal muscle satellite cells were cultured from mature rats and were treated in vitro with various combinations of transforming growth factor (TGF)-beta, fibroblast growth factor (FGF), and insulin-like growth factor I (IGF-I). In serum-free defined medium the following observations were made: TGF-beta depressed proliferation and inhibited differentiation; FGF stimulated proliferation and depressed differentiation; IGF-I stimulated proliferation to a small degree but demonstrated a more pronounced stimulation of differentiation. In evaluating combinations of these three factors, the differentiation inhibiting effect of TGF-beta could not be counteracted by any combination of IGF-I or FGF. The proliferation-depressing activity of TGF-beta, however, could not inhibit the mitogenic activity of FGF. Maximum stimulation of proliferation was observed in the presence of both FGF and IGF-I. The highest percentage fusion was also observed under these conditions, but differentiation with minimal proliferation resulted from treatment with IGF-I, alone. By altering the concentrations of TGF-beta, FGF, and IGF-I, satellite cells can be induced to proliferate, differentiate, or to remain quiescent.     

Interferon-gamma and interleukin-1 beta inhibit adipoconversion in cultured rodent preadipocytes.

Cytokines like tumor necrosis factor (TNF), interferon-gamma (IFN-gamma), and interleukin-1 (IL-1) are known to interfere with the differentiation of cultured cell lines of adipocyte precursors. In the present study, the effect of mouse and rat IFN-gamma, as well as human IL-1 beta, was investigated on rodent preadipocytes in primary cultures, either in the presence of fetal bovine serum (FBS, 10%) or in serum-free defined medium. IFN-gamma exerted an antiproliferative action that was more pronounced when cells reached confluency than during the growth phase of the culture. Morphological observation and quantifications of undifferentiated and differentiating cells revealed that IFN-gamma caused a decrease in the proportion of cells devoid of lipid droplets which would correspond to fibroblast-like cells, whereas preadipocytes remained unaffected. IFN-gamma induced a marked retardation of adipoconversion, resulting in a partial inhibition of lipoprotein lipase (LPL) activity and a severe decrease in glycerol-3-phosphate dehydrogenase (GPDH) activity. The antiproliferative and anti-LPL effects of IFN-gamma were neutralized by adding anti-IFN-gamma antibodies, while these antibodies prevented only partially the depressing effect of IFN-gamma on GPDH activity. Contrary to IFN-gamma, IL-1 beta slightly enhanced the proliferation in preadipocyte cultures. IL-1 beta also depressed adipoconversion, inhibited markedly LPL activity, and partially reduced GPDH activity. These results show that the influence of cytokines on adipoconversion observed in preadipocyte cell lines can be found in normal preadipocytes in culture.