No abstract

No abstract

Sterol response element binding protein (SREBP) is a key transcription factor in insulin and glucose metabolism. We previously demonstrated that elevated levels of membrane sphingomyelin (SM) were related to peroxisome proliferator–activated receptor-γ (PPARγ), which is a known target gene of SREBP-1 in adipocytes. However, the role of SM in SREBP expression in adipocytes remains unknown. In human abdominal adipose tissue from obese women with various concentrations of fasting plasma insulin, SREBP-1 proteins decreased in parallel with increases in membrane SM levels. An inverse correlation was found between the membrane SM content and the levels of SREBP-1c/ERK/Ras/PPARγ/CREB proteins. For the first time, we demonstrate the effects of SM and its signaling pathway in 3T3-F442A adipocytes. These cells were enriched or unenriched with SM in a range of concentrations similar to those observed in obese subjects by adding exogenous natural SMs (having different acyl chain lengths) or by inhibiting neutral sphingomyelinase. SM accumulated in caveolae of the plasma membrane within 24 h and then in the intracellular space. SM enrichment decreased SREBP-1 through the inhibition of extracellular signal-regulated protein kinase (ERK) but not JNK or p38 mitogen-activated protein kinase (MAPK). Ras/Raf-1/MEK1/2 and KSR proteins, which are upstream mediators of ERK, were down-regulated, whereas SREBP-2/caveolin and cholesterol were up-regulated. In SM-unmodulated adipocytes treated with DL-1-Phenyl-2-Palmitoylamino-3-morpholino-1-propanol (PPMP), where the ceramide level increased, the expression levels of SREBPs and ERK were modulated in an opposite direction relative to the SM-enriched cells. SM inhibited the insulin-induced expression of SREBP-1. Rosiglitazone, which is an anti-diabetic agent and potent activator of PPARγ, reversed the effects of SM on SREBP-1, PPARγ and CREB. Taken together, these findings provide novel insights indicating that excess membrane SM might be critical for regulating SREBPs in adipocytes via a MAPK-dependent pathway.

We have shown that membrane sphingomyelin (SM) is an independent predictor of the variance of fasting plasma insulin (FPI) concentrations and the homeostasis model assessment (HOMA) estimate of insulin resistance in obese women. The peroxisome proliferator-activated receptor-gamma (PPAR-gamma) is a key component in adipocyte differentiation that may also contribute to the sensitivity of cells to insulin. PPAR-gamma is activated by fatty acids, and the membrane composition may have an impact on the activity of PPAR-gamma and thus on the sensitivity of adipocytes to insulin. We investigated these possible links by determining the phospholipid contents of adipocyte membranes, the mRNA expression of PPAR-gamma, and the FPI and HOMA estimate of insulin resistance in obese women. The mRNA levels of tumor necrosis factor-alpha (TNF-alpha), which is suspected to play a role in insulin resistance and which downregulates PPAR-gamma expression, were also quantified. FPI and HOMA were strongly positively correlated with membrane SM (P < 0.005) and cholesterol (P < 0.005). PPAR-gamma mRNA levels were negatively correlated with FPI (P < 0.05) and HOMA (P < 0.05) and positively correlated with high-density lipoprotein (HDL) cholesterol (P < 0.05), membrane SM (P < 0.05), and cholesterol contents (P < 0.05). TNF-alpha mRNA levels were not correlated with membrane parameters. In stepwise multiple regression analysis, the variations in PPAR-gamma mRNA levels were mainly explained by HDL cholesterol (31.9%) and membrane SM (17.7%). Our study shows that the expression of PPAR-gamma, a major factor controlling adipocyte functions, the lipid composition of the membrane, and insulin sensitivity are probably closely associated in the adipose tissue of obese women.

BACKGROUND: The cell functions involved in the action of insulin Ð receptor binding, enzyme and transporter activities Ð are controlled by membrane properties. We have previously shown that the fasting plasma insulin (FPI) concentration and the homeostasis model assessment (HOMA) estimate of insulin resistance are associated with the sphingomyelin concentration in the erythrocyte membranes of obese women. OBJECTIVES: (1) To study the distribution of phospholipid classes in the plasma membrane and their association with insulin resistance markers in the adipocyte, an insulin-sensitive cell in obese women. (2) To investigate the in¯uence of diabetes in a small group of obese women treated by diet alone. (3) To compare the distribution of phospholipids in erythrocyte membranes in a subgroup of obese nondiabetic and diabetic women. SUBJECTS: Subcutaneous fat biopsies were taken from the abdominal region of 19 obese non-diabetic and seven obese type 2 diabetic women. Erythrocyte membrane assessment was performed in a subgroup of 10 of the 19 obese nondiabetic and in the seven diabetic patients. METHODS: The phospholipid composition of adipocyte and erythrocyte plasma membranes was analyzed by high performance liquid chromatography. RESULTS: FPI was positively correlated with the adipocyte membrane contents of sphingomyelin (P`0.001), phosphatidylethanolamine (P`0.05), and phosphatidylcholine (P`0.01) in the obese nondiabetic women. Similar correlations were obtained with HOMA. A stepwise multiple regression analysis indicated that sphingomyelin accounted for 45.6 and 43.8% of the variance in FPI and HOMA values as an independent predictor. There was a similar positive independent association between FPI and SM in the erythrocyte membranes of the studied subgroup. Diabetes per se did not in¯uence the independent association between SM membrane contents and FPI in both cell types. CONCLUSION: These results suggest a link between membrane phospholipid composition, especially SM, and hyperinsulinemia in obese women. International Journal of Obesity (2000) 24, 1600±1607 Keywords: obesity; type 2 diabetes; hyperinsulinemia; adipocyte; erythrocyte; plasma membrane; phospholipid classes IntroductionLow insulin sensitivity is common in obese individuals and is primarily due to defects in the pathway of insulin action in target tissues: muscle, liver and adipose tissue. It has been suggested recently that insulin resistance is associated with alterations in cell membrane properties. 1,2 Recent studies have demonstrated relationships between the fatty acid composition of phospholipids in skeletal muscle and the action of insulin. 3 ± 8 We have previously shown that changes in the distribution of phospholipid classes of the erythrocyte membrane were also associated with fasting plasma insulin (FPI) concentration and the homeostasis model assessment (HOMA) estimate of insulin resistance in 69 obese nondiabetic normoglycemic women. The most insulin resistant patients had the highest erythrocyte membrane sphingomyelin content....

No abstract

The membrane fluidity of reduced-amphotericin B (AmB)-sensitivity Kluyveromyces lactis mutant strain is higher than that of the wild-type K. lactis strain. After culture of the K. lactis and K. lactis mutant cells in the presence of subinhibitory doses of AmB (10 and 125 mg/liter, respectively), the plasma membranes of both yeast strains also showed a higher fluidity than did those of control cells. High membrane fluidity was associated with changes in the structural properties of the membranes. Culture of the K. lactis and K. lactis mutant cells in the presence of AmB induced changes in membrane lipid contents. In particular, phospholipid contents were increased in both strains treated with AmB, compared with their corresponding counterparts. As a result, the sterol/phospholipid ratio decreased. The relative proportion of monounsaturated fatty acids also increased after AmB treatment. The saturated fatty acid/monounsaturated fatty acid ratio decreased in K. lactis and K. lactis mutant cells treated with AmB but also in K. lactis mutant control cells compared to that in the K. lactis wild strain. These changes in lipid composition explain the higher fluidity, which could represent a process of metabolic resistance of the yeasts to AmB.Antifungal heptaen amphotericin B (AmB) is presently considered the "gold standard" for the intravenous chemotherapy of deep fungal infections in spite of its secondary effects (24). This antibiotic has been a therapeutic agent since 1956, but its mode of action at the molecular level has not been totally elucidated. Various interaction models with membrane sterols have been proposed. However, the most recent models are based on the formation of stable complexes not only with sterols but also with membrane phospholipids whose interactions induce pore formation (1, 6).AmB was isolated from Streptomyces nodosus cultures by Gold et al. (9). It is a polyene belonging to the macrolide family, and its molecular conformation is closed to phospholipid molecules. This antifungal agent possesses a hydrophobic part (hydrocarbon chain) and a hydrophilic part (polyhydroxyl chain); its amphipathic properties allow interactions with the membrane after diffusion through the yeast cell wall. Its interaction with membrane constituents (14,16,18) provokes the formation of pores responsible for several reactions, such as lipid oxidations and peroxidations, inhibition of membrane enzymes (3, 4), osmotic shocks, and, finally, cell death (32).Yeast resistance to antifungal agents, mainly azol derivatives, is a well-known phenomenon. Yeast cells with reduced sensitivity to AmB have been isolated from patients (24, 29). Horsburgh et al. (13) and Hakkou et al. (11) also isolated a Candida lusitaniae strain and a Kluyveromyces lactis strain, respectively, with reduced sensitivity to AmB. The reduced sensitivity of the strains either may stem from changes in the membrane chemical properties which may result from mutations or may be induced by the antifungal agent itself. The alterations of the membrane may al...

We recently demonstrated that the sphingomyelin (SM) content of adipocyte membranes was negatively correlated with the expression of peroxisome proliferator-activated receptor-Q Q (PPARQ Q) in the subcutaneous adipose tissue of obese women with variable degrees of insulin resistance. We have now investigated whether SM really does have an impact on the expression of PPARQ Q in 3T3-F442A adipocytes. Adding SM to the culture medium for 24 h caused a significant increase in SM content of adipocyte membranes and an acyl chain lengthdependent decrease in the levels of PPARQ Q mRNA and protein.The longer the acyl chain of the fatty acid of SM, the greater was the decrease in PPARQ Q. These data suggest that the nature of the fatty acid is important in the regulation of PPARQ Q by the SM pathway. ß