The effect of pregnancy on the control of lipolysis in fat cells isolated from human adipose tissue.

1. Lipolysis has been estimated by measuring the release of glycerol in isolated adipose tissue cells obtained from women in early prognancy, late pregnancy and 1 - 3 days post partum and from non-pregnant women. 2. Adipocytes of women at the end of pregnancy exhibited higher rates of lipolysis in response to adrenaline (1.5 - 15 muM) plus phentolamine (13 muM) than those of non-pregnant women or those in early pregnancy. 3. Lipolysis in response to adrenaline plus phentolamine in fat cells from women 1 - 3 days post partum was reduced compared to that at the end of gestation but was enhanced relative to that in the non-pregnant or early pregnant state. 4. Basal lipolysis also tended to be greatest at term. 5. Under conditions where the production of cyclic AMP was not rate limiting for the stimulation of lipolysis, that is in the presence of dibutyryl cyclic AMP (1 mM) or adrenaline (15 muM) plus phentolamine (13 muM) plus caffeine (1 mM), the release of glycerol in cells from women at term and in the puerperium was greater than that in women in the non-pregnant or early pregnant state. 6. Cell levels of cyclic AMP rose after incubation with adrenaline (6 muM) plus phentolamine or adrenaline (15 muM) plus phentolamine plus caffeine (1 mM) but were similar in all four groups of women. 7. It is concluded that the observed enhancement of lipolysis demonstrated in fat cells from women at the end of pregnancy reflects an increase in hormone-sensitive lipase activity rather than a modification of hormone receptor site sensitivity or of the rates of synthesis or breakdown of cyclic AMP. 8. This increase in adipose tissue lipolysis at the end of gestation could contribute to the reported rise in plasma nonesterified fatty acids in the final weeks of pregnancy.     

Human Adipose Tissue in Culture

Explants of human adipose tissue were cultured for several days with noradrenaline (NA). After the culture period the explants were carefully washed, preincubated and then incubated for 2 h by a technique analogous to those generally used to study adipose tissue metabolism. The results show that explants cultured with NA were considerably less responsive to the acute lipolytic effect of catecholamines than explants not previously exposed to NA. The diminished responsiveness could not be reversed by the addition of phentolamine or by preincubating the explants with a prostaglandin antagonist and it was dependent upon the concentration of noradrenaline used in the culture medium as well as upon the period of time that the explants were exposed to the hormone. The cyclic AMP (cAMP) levels did not increase in response to NA in these explants. Addition of theophylline or dibutyryl cAMP elicited a lipolytic response. However, the phosphodiesterase levels were not increased in the explants cultured with noradrenaline. When freshly excised tissue specimens were incubated in the culture medium which had previously contained NA the acute lipolytic effect of catecholamines was blunted. The results indicate that an antagonist is formed during the incubation with NA which inhibits the responsiveness to catecholamines. The effect of the antagonist seems to be exerted at a step prior to the formation of cAMP.     

The Composition and Metabolism in Vitro of Adipose Tissue Fat Cells of Different Sizes

Abstract. Fat cells of different sizes were isolated from the same sample of adipose tissue by collagenase treatment and fractionation by flotation in an isoosmolar medium. Protein, phospholipid and cholesterol increased with increasing fat cell size. Cholesterol correlated most closely with fat cell volume but phospholipid apparently with surface. Most of fat cell cholesterol was found in the triglyceride droplet, and only a smaller part in a particulate fraction from fat cells. Incorporation of labelled glucose into triglyceride increased with fat cell size, apparently in proportion to fat cell surface rather than diameter or volume. This was the case also with incorporation into fatty acids. The association between fat cell size and metabolism was present also in alloxan diabetic rats. Short term insulin deficiency thus does not abolish the increase of metabolic activity with fat cell size. Analyses of activities of selected glycolytic enzymes and of fatty acid synthesis from acetate in cell-free systems showed a similar dependence on fat cell size, demonstrating that isotope dilution phenomena in large and small fat cells are probably not responsible. — Lipolytic activity in the basal state and after epinephrine stimulation was increased in large fat cells, which also reesterified more fatty acids than small fat cells when calculated according to the balance method. — It was concluded that larger fat cells are metabolically more active than smaller fat cells. The increase of glyceride-glycerol labelling and of glycerol release from larger fat cells suggests an increased triglyceride turn-over in these fat cells. Large fat cells might thus be considered as an active metabolic sub-compartment of adipose tissue.     

Differences in Lipolysis between Human Subcutaneous and Omental Adipose Tissues

Hydrolysis of triglycerides to fatty acids and glycerol in fat cells (lipolysis) is of importance for the control of lipid and carbohydrate metabolism. This process is regulated by several hormones and parahormones acting on cyclic AMP formation or breakdown, which in turn influences the activity of hormone sensitive lipase. The latter enzyme stimulates hydrolysis of triglycerides in fat cells. It is well established through in vivo and in vitro investigations that there are regional variations in the lipolytic activity of human adipose tissue. The rate of lipolysis is low in the subcutaneous femoral/gluteal region, intermediate in the subcutaneous abdominal region and high in the visceral (i.e. omental) region. In non-obese subjects the differences between the subcutaneous and visceral fat depots may be explained by site variations in the function of receptors for insulin, catecholamines and adenosine. The lipolytic beta₁ and beta₂ adrenoceptors, as well as the newly discovered beta₃, are most active in the visceral fat cells. The antilipolytic insulin receptors, alpha₂ adrenoceptors and adenosine receptors are most active in the subcutaneous fat cells. In subjects with upper-body obesity the regional variations in the action of catecholamines on lipolysis are further enhanced. Decreased action of beta₂-adrenergic receptors and increased activity of alpha₂-adrenergic adrenoceptors in combination with defects in hormone sensitive lipase function inhibits the lipolytic effect of catecholamines in subcutaneous fat cells whereas increased activity of beta₃-adrenergic receptors and decreased activity of alpha₂ adrenoceptors augment the lipolytic response in visceral fat cells. These abnormalities in catecholamine function promote release of free fatty acids from the visceral fat cells to the liver through the portal system and might cause several of the metabolic complications to upper-body obesity.     

Increased plasma cyclic AMP concentrations in fasting man

Plasma concentrations of adenosine 3′, 5′ monophosphate (cyclic AMP), free fatty acids (FFA), glycerol and glucose were measured after 24, 48 and 72 h fasting in healthy male students. There was a marked rise in plasma concentrations of cyclic AMP, FFA and glycerol from 24 to 48 h fasting with no further changes from 48 to 72 h fasting. Plasma glucose concentrations fell. After 72 h fasting, inhibition of adipose tissue lipolysis with a nicotinic acid analogue effected no significant change in plasma cyclic AMP concentrations, indicating that the elevated plasma nucleotide concentrations during fasting were not related to stimulation of lipolysis.     

The Effects of Starvation on Lipogenesis in Human Adipose Tissue

Abstract 1. The effects of starvation on glyceride synthesis have been studied in biopsies of human adipose tissue. After a week's fast glyceride synthesis from glucose (8.3 and 14 mM) was reduced by about 70% in intact adipose tissue; this occurred within 5 days of starting the fast.—2. A method has been devised to follow the simultaneous incorporation of ¹⁴C-l-palmitate into acyl-CoA, phospholipid and neutral lipid in homogenates of human adipose tissue. This system was sensitive to fluoride which produced an accumulation of radioactivity in phospholipid and a reduction in neutral lipid.— 3. Starvation also reduced the incorporation of ¹⁴C-l-palmitate into neutral lipid in a similar incubation system, but radio-activity now accumulated in long chain acyl-CoA. This suggests that fasting is accompanied by an inhibition of L-glycerol 3-phosphate acyl transferase (E.C. 2.3.1.15).—4. Oleate (600 μM), which is similar to concentrations found in homogenates of fasting adipose tissue, was found to inhibit the activity of acyl transferase by about 80%. The possibility was therefore considered that the reduction of lipogenesis during fasting is mediated by free fatty acid inhibiting the acyl transferase.     

Methodological Aspects of Protein-Binding Assays for Cyclic AMP in Human Adipose Tissue

The specificity of the protein-binding assay for cyclic AMP according to Gilman's method (11) was studied in human adipose tissue. A kit assay obtained from Boehringer-Mannheim was used for the study. Impurities in the solvents used for homogenization and extraction influenced the assay and led to serious overestimation of the actual amount of cAMP. This was corrected for by treating the standard and tissue samples in the same manner. Extracts from human adipose tissue contained material that increased the binding of cAMP to the binding protein and led to underestimation of intracellular cAMP by 60–70 per cent. This was overcome by using a protein kinase inhibitor in the assay. Furthermore, when human adipose tissue was incubated in an albumin-containing medium, intracellular cAMP was underestimated by 30 per cent, since albumin increased the binding of cAMP to the binding protein.     

胎球蛋白-a对3T3-L1脂肪细胞脂肪分解的影响

目的观察胎球蛋白-a(fetuin-a)对脂肪分解的影响并探讨其可能的机制。方法体外培养3T3-L1前脂肪细胞,分化成熟后用不同浓度的fetuin-a孵育。采用甘油检测试剂盒测定释放到上清液的甘油含量;采用Real-timePCR检测细胞中脂肪甘油三酯脂肪酶(Adipose triglyceride lipase,ATGL)的mRNA表达;采用western blot检测细胞内磷酸化激素敏感脂肪酶(Hormone Sensitive Lipase,HSL)和ATGL的蛋白表达;采用ELISA试剂盒检测细胞内环磷酸腺苷(cyclic adenosine monophosphate,cAMP)的含量。结果 Fetuin-a能够抑制成熟脂肪细胞的脂肪分解,具有剂量依赖关系。Fetuin-a能够降低细胞内cAMP及磷酸化的HSL含量,同时使脂肪细胞ATGL mRNA水平及蛋白表达降低。结论 Fetuin-a抑制3T3-L1脂肪细胞的脂肪分解,其机制可能是通过抑制cAMP-PKA-HSL通路以及降低细胞内ATGL的表达而发挥作用的。     

The Influence of Body Weight and Cell Size on Lipogenesis and Lipolysis of Isolated Rat Fat Cells*

Abstract. A method for fractionating isolated fat cells is described. After fixation with osmium tetroxide, the cells are passed through a series of polyamide filters of decreasing mesh width and the number of cells on each filter is counted. – Isolated fat cells from ad libitum fed rats weighing 85–580 g were incubated with [U-¹⁴C]-glucose (0.55 mM), fixed with osmium tetroxide and fractionated according to size. When the cells were obtained from one animal the conversion of glucose to neutral lipids per cell increased with increasing cell size just as well in the absence of insulin as in the presence of insulin (10 mU/ml). The following results were obtained from comparisons between animals of different weight: in the absence of insulin, the lipogenesis for cells of the same size remained constant irrespective of the rat weight, whereas in the presence of insulin the lipogenesis for cells of the same size decreased markedly when the rat weight exceeded about 300 g. It is concluded that factors other than the cell size per st (e.g. age, degree of obesity) determine the responsiveness to insulin. – The hormone-stimulated lipolysis was studied on unfractionated cell samples from ad libitum fed rats of different weights and the following was found: the glycerol release per 10⁵ cells induced by ACTH and norepinephrine in maximally stimulating concentrations increased with increasing mean cell size or rat weight. In contrast, the glucagon-induced lipolysis of cells from rats weighing 300–400 g was smaller than that of cells from rats weighing 100–120 g.     

The Effect of Adult Diabetes on Glucose Utilization and Esterification of Palmitate by Human Adipose Tissue

Abstract. 1. The conversion of glucose and palmitate to neutral lipid by adipose tissue has been studied in a group of 30 adult diabetics and compared to a group of 30 non-diabetic controls. — 2. Adipose tissue from both sources showed similar saturation characteristics for glucose; uptake of palmitate was linear up to 3 mM. — 3. Adipose tissue of diabetics incubated with glucose (14 mM) converted 270 ± 60 (90) mμmoles/mg protein/2 hours to neutral lipid compared to non-diabetics who converted 380 ±70 (10) mμmoles/mg protein/2 hours. Similar results were obtained with a medium glucose concentration of 8 mM. No differences were found in glucose utilization between a group of 7 lean diabetics and 6 obese diabetics. — 4. Conversion of palmitate to neutral lipid by adipose tissue was inhibited by cyanide (1.5 mM), fluoride (50 mM) and was found to be temperature dependent. — 5. Adult diabetics converted 0.36 ± 0.06 (17) mμmoles palmitate/mg protein/2 hours into neutral lipid, compared to a group of non-diabetics who converted 1.09 ± 0.35 (13) mμmoles/mg protein/2 hours.— 6. The significance of these observations is discussed in relation to the impaired glucose tolerance test of diabetes.     

The Glucose Uptake of Human Adipose Tissue in Obesity

Abstract. Radioactivity in adipose tissue triglycerides was followed after giving an intravenous injection of 25 g of glucose with 50 μCi of U-¹⁴C-glucose to 12 obese and 7 control patients. Seventy-five minutes after injection the radioactivity could be measured with sufficient precision. Recirculation of label was probably small during this period. Total adipose tissue glucose uptake could therefore be estimated knowing the variation of adipose tissue triglyceride radioactivity in different selected regions, and total body fat, and was found to be only a few per cent of all assimilated glucose in the body. Triglyceride radio-activity per gram of adipose tissue was similar in obese subjects and controls. Consequently total adipose tissue label was higher in the obese. Triglyceride radioactivity per fat cell correlated positively with fat cell size which in turn correlated positively with plasma insulin concentration. During the two first weeks after injection adipose tissue triglyceride radioactivity increased after which it began to fall in some patients.     

Insulin resistance, lipid and fatty acid concentrations in 867 healthy Europeans. European Group for the Study of Insulin Resistance (EGIR)

BACKGROUND: Insulin resistance, dyslipidaemia and abnormal nonesterified fatty acid (NEFA) metabolism are features of the 'metabolic syndrome', but the mechanisms of these relationships are uncertain. MATERIALS AND METHODS: We studied associations between insulin resistance and lipoprotein concentrations by retrospective analysis of euglycaemic hyperinsulinaemic clamp data from 867 normoglycaemic subjects in 21 European centres. Data on NEFA concentrations were available in a subgroup of 541 subjects from 9 clinical centres. These subjects' characteristics do not vary significantly from those of the whole cohort. RESULTS: After adjustment for the effects of age, sex, obesity and intercentre variability, regression analysis showed relationships between triglycerides and markers of insulin sensitivity. There were significant correlations between triglycerides and fasting plasma glucose (P < 0.0001), fasting plasma insulin (P < 0.0001) and mean glucose infusion rate at steady state (M-value, P < 0.0001). Indices of insulin resistance were related to NEFA concentrations. Fasting NEFA were negatively correlated with the M-value (P < 0.0001). Non-esterified fatty acids at steady state were positively correlated with fasting markers of insulin resistance: fasting plasma glucose (P < 0.05), fasting plasma insulin (P < 0.005) and negatively correlated with the M-value (P < 0.0005). There were relationships between fasting concentrations of plasma lipids and of NEFAs. Non-esterified fatty acids at steady state correlated with fasting triglycerides (P < 0.0001), but not with any of the other plasma lipoprotein concentrations. The associations of fasting triglycerides with the M-value and with NEFAs at steady state were independent of each other. All these associations were independent of obesity and geographical location CONCLUSION: The results in this large cohort of healthy European subjects suggest that triglyceride concentrations depend upon both insulin's gluco-regulation (estimated by glucose uptake) and antilipolytic insulin action (measured by NEFA levels) during an euglycaemic clamp.     

不同类型冠心病患者血浆游离脂肪酸水平变化及其与病变程度的关系

目的探讨血浆游离脂肪酸(NEFA)水平变化与冠心病(CAD)病变程度的相关性。方法选取疑为CAD患者84例,分别行生化检验和冠状动脉造影检查;根据冠状动脉造影结果将其分为4组,分别为不稳定型心绞痛组(n=46)、稳定型心绞痛组(n=9)、急性心肌梗死组(n=20)、阴性对照组(n=9),对比组间血脂指标与NEFA是否存在差异分析,CAD患者发生急性心肌梗死风险与造影特点。结果 CAD组NEFA水平均高于阴性对照组,差异有统计学意义(P0.05);急性心肌梗死组NEFA水平高于稳定型心绞痛组和不稳定型心绞痛组,差异有统计学意义(P0.05);Logistic回归分析显示,校正后NEFA≥0.415mmol/L组出现急性心肌梗死的风险是NEFA0.415mmol/L组的2.61倍,95%CI:1.33~5.02,P0.05;不同水平的NEFA患者GENSINI积分、狭窄冠状动脉支数与3支冠状动脉病变数量差异无统计学意义(P0.05)。结论 CAD患者血浆NEFA出现升高,急性心肌梗死患者升高明显,但其与病变程度相关性未明确。     

The Sources of Plasma Cyclic AMP: Studies in the Rat using Isoprenaline, Nicotinic Acid and Glucagon

Abstract. The effects of intravenous administration of isoprenaline, glucagon and nicotinic acid on plasma concentrations of cyclic AMP in rats are described. In order to determine the relative importance of the liver as a source of extracellular cyclic AMP, the effects of the hormones were investigated in intact and functionally hepatectomised rats. The results showed that hepatectomy did not prevent an isoprenaline-stimulated increase in plasma cyclic AMP concentrations, although glucagon was without effect on plasma nucleotide concentrations in this group of animals. It is suggested that the liver is essential for the action of glucagon but that isoprenaline can increase plasma cyclic AMP concentrations in hepatectomised animals by increasing extrahepatic release of the nucleotide. Since inhibition of adipose tissue lipolysis with nicotinic acid did not prevent an isoprenaline or glucagon-stimulated increase in plasma cyclic AMP concentrations, adipose tissue is discounted as a major source of plasma cyclic AMP.     

The sources of plasma cyclic AMP: studies in the rat using isoprenaline, nicotinic acid and glucagon.

The effects of intravenous administration of isoprenaline, glucagon and nicotinic acid on plasma concentrations of cyclic AMP in rats are described. In order to determine the relative importance of the liver as a source of extracellular cyclic AMP, the effects of the hormones were investigated in intact and functionally hepatectomised rats. The results showed that hepatectomy did not prevent an isoprenaline-stimulated increase in plasma cyclic AMP concentrations, although glucagon was without effect on plasma nucleotide concentrations in this group of animals. It is suggested that the liver is essential for the action of glucagon but that isoprenaline can increase plasma cyclic AMP concentrations in hepatectomised animals by increasing extrahepatic release of the nucleotide. Since inhibition of adipose tissue lipolysis with nicotinic acid did not prevent an isoprenaline or glucagon-stimulated increase in plasma cyclic AMP concentrations, adipose tissue is discounted as a major source of plasma cyclic AMP.     

超声波对早孕蜕膜中免疫活性细胞影响的研究

采用诊断超声强度经腹部辐照宫腔.取早孕蜕膜,检测其中的免疫活性细胞,以探讨超声波对早孕蜕膜中免疫活性细胞的影响。结果表明:诊断超声强度未能影响早孕蜕膜中T淋巴细胞及其亚群的状态,却可使巨噬细胞数目减少,或许在一定程度上会削弱宫腔局部的抗感染能力。     

妊娠晚期孕妇血hCG、HPL及hCG/HPL异常与不良妊娠结局关系的研究

目的:通过对妊娠晚期孕妇进行人绒毛膜促性腺激素(hCG)、胎盘泌乳素(HPL)及二者比值的测定,探讨其与不良妊娠结局的关系,以指导临床治疗。方法:随机选取住院分娩孕妇565例,采用放射免疫RIA法进行hCG、HPL检测,并进行统计学处理。结果:早产、胎膜早破及妊娠期高血压综合征患者血清hCG水平明显高于正常妊娠患者(P<0.05,P<0.01),而不良妊娠结局患者与正常妊娠患者HPL检测结果比较无显著差异(P>0.05),但hCG/HPL比较差异非常显著(P<0.01)。结论:hCG可作为预测不良妊娠结局的检测指标之一,监测hCG/HPL可提高高危妊娠的检出率。     

罗格列酮对2型糖尿病游离脂肪酸的影响

目的观察罗格列酮对2型糖尿病游离脂肪酸（FFA）的影响。方法60例2型糖尿病患者,随机分为罗格列酮组与二甲双胍组。治疗前和治疗3个月后,测量身高和体质量,计算体质量指数（BMI）,观察空腹血糖（FBS）、甘油三酯（TG）、总胆固醇（TC）、高密度脂蛋白胆固醇（HDL-C）、低密度脂蛋白胆固醇（LDL-C）、FFA、糖化血红蛋白（HbA1c）的变化。结果治疗后两组FBS、HbA1c均下降。罗格列酮组FBS治疗前后（10．34±3．08）mmol／L vs（7．75±1．46）mmol／L（P〈0．01）,HbA1c（8．85±2．28）％vs（7．28±1．33）％（P〈0．01）;二甲双胍组FBS（9．51±2．89）mmol／Lvs（7．36±1．40）mmol／L（P〈0．01）,HbA1c（9．26±2．45）％vs（7．61±1．23）％（P〈0．01）。但两组之间比较无统计学意义（P〉0．05）。罗格列酮组治疗前后FFA（0．65±0．24）mmol／Lvs（0．54±0．19）mmol／L（P〈0．05）。而二甲双胍组治疗前后FFA无明显下降。结论罗格列酮治疗降低FFA的水平,有助于减少2型糖尿病大血管并发症的发生。     

骨髓、脐带及脂肪来源间充质干细胞的成脂能力存在差异

间充质干细胞（mesenchymalstemcells,MSC）的来源非常广泛。由于不同组织来源的影响,MSC之间都会存在一定的差异。本研究分析比较脐带、脂肪及骨髓组织来源的MSC的基本生物学特性。从脐带、脂肪和骨髓分离培养MSC,在显微镜下观察这3种来源的MSC的形态（UC—MSC,AD—MSC和BM—MSC）,用流式细胞术诱导分化试验及定量荧光PCR分别检测UC-MSC,AD-MSC和BM-MSC的免疫表型、分化能力和过氧化物酶增殖激活受体-γ（peroxisomeproliferator—activatedreceptor-1,PPAR-1）mRNA的表达水平。结果表明,UC-MSC、AD-MSC和BM—MSC的形态都是成纤维细胞样;经罗丹明和DAPI染色后,用共聚焦显微镜观察发现它们的细胞形态类似;这三种来源Msc的免疫表型符合Msc的鉴定标准,而且表达水平一致;这三种来源MSC的成骨分化潜能相似,而成脂分化潜能存在较明显的差异,其中AD—MSC成脂能力最强,BM-MSC次之,UC-MSC最差。检测脂肪形成早期起重要作用的过氧化物酶增殖激活受体-γ（PPAR-1）mRNA表达水平在三种来源MSC中的基础表达水平,发现PPAR-γ mRNA在AD-MSC中最高,在BM—MSC次之,而在UC—MSC中最低,与成脂能力的表现相一致。这表明三种来源MSC成脂能力存在差异可能与它们PPAR-1的基础表达水平有关。结论：UC—MSC,AD—MSC和BM—MSC的形态类似;免疫表型符合MSC鉴定标准,表达水平一致;成骨分化能力相似,而成脂分化能力不同;PPAR-γ mRNA表达水平不一。关于差异的相关机制有待进一步研究。