Hyperthermia effect on lipolytic processes in rat blood and adipose tissue

Rats anaesthetized with Brevinarcon were placed in a high-temperature chamber (air temperature 50 degrees C, relative humidity 50%) for induction of hyperthermia (rectal temperature 41.0 +/- +/- 0.5 degrees C). The control group comprised rats anaesthetized in the same way but kept at room temperature. In the serum in both groups glucose, free fatty acids, immunoreactive insulin, lipolytic activity and ability to mobilize lipids in vitro were determined. It was shown that the glucose and free fatty acid levels and the activity mobilizing serum lipids in vitro in the rats subjected to hyperthermia were lower than in the control group by 12%, 23% and 150% respectively. The lipolytic activity of the serum of rats subjected to hyperthermia was 42% higher, and the level of immunoreactive insulin rose by about 224% in relation to the control group. These results point to inhibition of lipolysis in the adipose tissue with simultaneous activation of intravascular lipolysis during hyperthermia in rats.     

Bacteria-derived human growth hormone lacks lipolytic activity in rat adipose tissue

Bacteria-derived human growth hormone (hGH) shows little $\text{in}\text{vitro}$ lipolytic activity in adipose tissue from fed rats. In adipose tissue from fasted rats no lipolytic activity is observed. However, bacteria-derived hGH increased serum free fatty acids after intraperitoneal administration to hypophysectomized rats to the same extent as purified pituitary hGH. The dose response of the bacteria-derived hGH tested for $\text{in}\text{vitro}$ insulin-like activity was very similar to the pituitary extracted material. Thus bacteria-derived hGH behaves in a manner indistinguishable from highly purified preparations of pituitary hGH.     

Alkaline lipolytic activity of rabbit adipose tissue: Genotypes and their inheritance

A genetic variant of alkaline lipolytic activity (ALA), a soluble esterase found in mammalian adipose tissue, has previously been described. This slow variant was characterized by slower migration during starch gel electrophoresis, a decreased tendency to dimerize, and reduced enzymatic activity in constrast to the fast electrophoretic phenotype. The present paper identifies a homozygous (FF) and a heterozygous (Ff) fast phenotype which are differentiated after starch gel electrophoresis by the presence or absence of a slow band identical to that of the homozygous slow (ff)/it phenotype. In addition, ALA of homozygous and heterozygous fast rabbits differed in total enzymatic activity and its tendency to dimerize. The inheritance of this variant was proven to be Mendelian.This work was supported in part by United States Public Health Service Grants AM-06872 from the National Institute of Arthritis and Metabolic Diseases and GM-15874 from the National Institute of General Medical Sciences; and Project #417 of the Children's Bureau, Department of Health, Education and Welfare.     

Insulin inhibits lipolytic activity and degradation of beta-lipotropin in rabbit adipose tissue

beta-Lipotropin, a pituitary peptide, is a potent stimulator of lipolysis in rabbit adipose tissue in vitro and in vivo. Insulin inhibited the beta-lipotropin (1-100 nM)-stimulated glycerol release from rabbit adipocytes and fat pads significantly at concentrations of 10 and 100 microM. Both these concentrations of insulin also decreased the degradation of beta-lipotropin in intact adipose tissue to the same extent as the lipolytic activity. Furthermore, insulin reduced the degradation of beta-lipotropin in rabbit adipose tissue homogenate. Like insulin, several lysosomotropic agents also decreased significantly the degradation and the lipolytic activity of beta-lipotropin. On the other hand, insulin-like growth factor I in lower concentrations (1-100 nM) did not effect degradation and lipolytic activity of beta-lipotropin in rabbit adipose tissue. Thus, a direct influence of insulin on lysosomal enzymes degrading beta-lipotropin in rabbit adipose tissue can be suggested.     

Effect of hypothermia on lipolytic processes in blood and adipose tissue of rat

Short-lasting hypothermia raises the FFA level in the blood and this rise is associated with increased lipid-mobilizing activity and higher lipolytic activity of the serum. Raised FFA level and increased lipid-mobilizing activity of the serum persist even when the degree of general anaesthesia is sufficient for preventing thermogenesis signs (shivering and piloerection) caused by falling body temperature. Beta-adrenergic blockade fails to abolish the effect of lipolysis activation caused by hypothermia. These observations suggest that during hypothermia in the blood of the animals appear factors stimulating lipolysis in the adipose tissue. One of these factors may stimulate tissue lipolysis independently of beta-adrenergic receptors. Insulin blocks significantly lipolytic processes in the adipose tissue of hypothermic animals, but its administration is connected with the danger of hypoglycaemia development.     

Modulation of white adipose tissue proteome by aging and calorie restriction

Aging is associated with an accrual of body fat, progressive development of insulin resistance and other obesity comorbidities that contribute to decrease life span. Caloric restriction (CR), which primarily affects energy stores in adipose tissue, is known to extend life span and retard the aging process in animal models. In this study, a proteomic approach combining 2‐DE and MS was used to identify proteins modulated by aging and CR in rat white adipose tissue proteome. Proteomic analysis revealed 133 differentially expressed spots, 57 of which were unambiguously identified by MS. Although CR opposed part of the age‐associated protein expression patterns, many effects of CR were on proteins unaltered by age, suggesting that the effects of CR on adipose tissue are only weakly related to those of aging. Particularly, CR and aging altered glucose, intermediate and lipid metabolism, with CR enhancing the expression of enzymes involved in oxalacetate and NADPH production, lipid biosynthesis and lipolysis. Consistently, insulin‐β and β3‐adrenergic receptors were also increased by CR, which denotes improved sensitivity to lipogenic/lipolytic stimuli. Other beneficial outcomes of CR were an improvement in oxidative stress, preventing the age‐associated decrease in several antioxidant enzymes. Proteins involved in cytoskeleton, iron storage, energy metabolism and several proteins with novel or unknown functions in adipose tissue were also modulated by age and/or CR. Such orchestrated changes in expression of multiple proteins provide insights into the mechanism underlying CR effects, ultimately allowing the discovery of new markers of aging and targets for the development of CR‐mimetics.     

Thiourea enhances mapping of the proteome from murine white adipose tissue

Adipose tissue imposes problems in two-dimensional (2-D) analysis due to its extremely high content of fat. To improve protein separation detergents and chaotropes were varied in the IEF step. The most important factor for obtaining distinct spots in the 2-D gel was whether thiourea was included or not. Many high molecular weight spots became resolved by using thiourea, while no spots disappeared or showed inferior characteristics, thus approximately twice as many spots were possible to quantify. Hydrophobic indices were compared for a set of proteins that gave rise to sharper spots with proteins that were not improved on the use of thiourea. The comparison did not give any statistically significant difference between the two groups of proteins. One of the effects obtained by inclusion of thiourea was that the dominating protein, serum albumin, appeared as more condensed spots allowing other minor proteins to be detected. This work resulted in a protocol which greatly enhances the resolution of proteins in adipose tissue. A 2-D map of mouse white adipose tissue from epididymal fat pads was constructed in which 140 spots were identified by mass spectrometry. This work lays the ground for our further studies on white adipose tissue in metabolic diseases such as obesity and dyslipidemia.     

Amyloidogenic and associated proteins in systemic amyloidosis proteome of adipose tissue

In systemic amyloidoses, widespread deposition of protein as amyloid causes severe organ dysfunction. It is necessary to discriminate among the different forms of amyloid to design an appropriate therapeutic strategy. We developed a proteomics methodology utilizing two-dimensional polyacrylamide gel electrophoresis followed by matrix-assisted laser desorption/ionization mass spectrometry and peptide mass fingerprinting to directly characterize amyloid deposits in abdominal subcutaneous fat obtained by fine needle aspiration from patients diagnosed as having amyloidoses typed as immunoglobulin light chain or transthyretin. Striking differences in the two-dimensional gel proteomes of adipose tissue were observed between controls and patients and between the two types of patients with distinct, additional spots present in the patient specimens that could be assigned as the amyloidogenic proteins in full-length and truncated forms. In patients heterozygotic for transthyretin mutations, wild-type peptides and peptides containing amyloidogenic transthyretin variants were isolated in roughly equal amounts from the same protein spots, indicative of incorporation of both species into the deposits. Furthermore novel spots unrelated to the amyloidogenic proteins appeared in patient samples; some of these were identified as isoforms of serum amyloid P and apolipoprotein E, proteins that have been described previously to be associated with amyloid deposits. Finally changes in the normal expression pattern of resident adipose proteins, such as down-regulation of alphaB-crystallin, peroxiredoxin 6, and aldo-keto reductase I, were observed in apparent association with the presence of amyloid, although their levels did not strictly correlate with the grade of amyloid deposition. This proteomics approach not only provides a way to detect and unambiguously type the deposits in abdominal subcutaneous fat aspirates from patients with amyloidoses but it may also have the capability to generate new insights into the mechanism of the diseases by identifying novel proteins or protein post-translational modifications associated with amyloid infiltration.     

Insulin and lipolytic hormones stimulate the same phosphodiesterase isoform in rat adipose tissue

Insulin sensitive phosphodiesterase from rat adipocytes is found in particulate fractions. Solubilisation of the enzyme with triton X-100 yields a preparation containing more than one phosphodiesterase activity as judged by its rate of thermal denaturation at 45 degrees C and by its non-linear kinetic plots. Immunoprecipitation of solubilised activity with a polyclonal antiserum raised against purified insulin-sensitive rat liver phosphodiesterase selected a form of the enzyme which showed a single exponential decay of enzyme activity when heated at 45 degrees C and linear low Km kinetics. Treatment of adipocytes with insulin ACTH, glucagon or isoproterenol stimulated the low Km particulate phosphodiesterase. The hormonal activation was retained following solubilisation and was also seen when activity was immunoprecipitated. It is suggested that all four hormones activate the same form of phosphodiesterase.