Expression of Fibroblast Growth Factor-21 in Muscle Is Associated with Lipodystrophy,Insulin Resistance and Lipid Disturbances in Patients with HIV

BackgroundFibroblast growth factor (FGF)-21 is a novel regulator of glucose and lipid metabolism. Recently, increased FGF-21 mRNA expression in muscle was found in patients with type 2 diabetes, but the role for FGF-21 in muscle is not well understood. Patients with HIV-infection and lipodystrophy are characterised by various degree of lipid-driven insulin resistance. We hypothesized that muscle FGF-21 mRNA would be altered in HIV patients with lipodystrophy.DesignTwenty-five HIV-infected men with lipodystrophy (LD) and 15 age-matched healthy controls, received an oral glucose tolerance test and a euglycemic-hyperinsulinemic clamp (50 mU/m2/min) combined with 6,6-H₂ glucose infusion. Muscle biopsies were obtained and FGF-21 mRNA and glycogen synthase (GS) activity were measured.ResultsSubjects with HIV were insulin resistant compared with non-HIV subjects. Compared to controls, HIV subjects demonstrated a twofold increase of plasma FGF-21 from 70.4±56.8 pg/ml vs 109.1±71.8 pg/ml, respectively (p = 0.04) and an eight-fold increase in muscular FGF-21 mRNA expression (p = 0.001). Muscle FGF-21 mRNA correlated inversely with the rate of disappearance of glucose during insulin clamp (r = −0.54, p = 0.0009), and the GS fractional velocity in muscle (r = −0.39, p = 0.03), and directly with fasting insulin (r = 0.50, p = 0.0022), HOMA-IR (r = 0.47, p = 0.004), triglycerides (r = 0.60. P = 0.0001), waist-to-hip ratio (r = 0.51, p = 0.0001) and limb fat mass (−0.46, p = 0.004), but not to plasma FGF-21.ConclusionFGF-21 mRNA is increased in skeletal muscle in HIV patients and correlates to whole-body (primarily reflecting muscle) insulin resistance, but not to plasma FGF-21. Those findings add to the evidence that FGF-21 is a myokine and may suggest that muscle FGF-21 is working in a local manner.     

Insulin stimulates interleukin-6 and tumor necrosis factor-alpha gene expression in human subcutaneous adipose tissue

High circulating levels of interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-alpha) are found in patients with hyperinsulinemia. Insulin stimulates release of IL-6 from adipocyte cultures, and it stimulates IL-6 gene expression in insulin-resistant, but not control, rat skeletal muscle. In addition, TNF-alpha may be involved in the pathogenesis of insulin resistance. Therefore, we studied the effect of insulin on IL-6 and TNF-alpha gene expression in human skeletal muscle and adipose tissue. Nine healthy young volunteers participated in the study. They underwent a 6-h hyperinsulinemic euglycemic clamp at a fixed insulin infusion rate, with blood glucose clamped at fasting level. Blood samples drawn at 0, 1, 2, 3, 4, 5, and 6 h were analyzed for IL-6 and TNF-alpha. Muscle and fat biopsies, obtained at 0, 2, 4, and 6 h, were analyzed for IL-6 and TNF-alpha mRNA with real-time PCR. IL-6 mRNA increased 11-, 3-, and 5-fold at 2, 4, and 6 h, respectively, in adipose tissue (ANOVA P = 0.027), whereas there was no significant effect of insulin on skeletal muscles. Plasma IL-6 increased during insulin stimulation. TNF-alpha mRNA increased 2.4-, 1.4-, and 2.2-fold in adipose tissue (ANOVA P = 0.001) and decreased 0.74-, 0.64-, and 0.68-fold in muscle tissue (ANOVA P = 0.04). Plasma levels of TNF-alpha were constant. In conclusion, the finding that insulin stimulates IL-6 and TNF-alpha gene expression in adipose tissue only and inhibits the TNF-alpha production in skeletal muscles suggests a differential regulation of muscle- and adipose tissue-derived IL-6 and TNF-alpha.     

Fibroblast Growth Factor 21 (FGF-21) in Peritoneal Dialysis Patients: Natural History and Metabolic Implications

Background

Human fibroblast growth factor 21 (FGF-21) is an endocrine liver hormone that stimulates adipocyte glucose uptake independently of insulin, suppresses hepatic glucose production and is involved in the regulation of body fat. Peritoneal dialysis (PD) patients suffer potential interference with FGF-21 status with as yet unknown repercussions.

Objectives

The aim of this study was to define the natural history of FGF-21 in PD patients, to analyze its relationship with glucose homeostasis parameters and to study the influence of residual renal function and peritoneal functional parameters on FGF-21 levels and their variation over time.

Methods

We studied 48 patients with uremia undergoing PD. Plasma samples were routinely obtained from each patient at baseline and at 1, 2 and 3 years after starting PD therapy.

Results

Plasma FGF-21 levels substantially increased over the first year and were maintained at high levels during the remainder of the study period (253 pg/ml (59; 685) at baseline; 582 pg/ml (60.5–949) at first year and 647 pg/ml (120.5–1116.6) at third year) (p<0.01). We found a positive correlation between time on dialysis and FGF-21 levels (p<0.001), and also, those patients with residual renal function (RRF) had significantly lower levels of FGF-21 than those without RRF (ρ -0.484, p<0.05). Lastly, there was also a significant association between FGF-21 levels and peritoneal protein losses (PPL), independent of the time on dialysis (ρ 0.410, p<0.05).

Conclusion

Our study shows that FGF-21 plasma levels in incident PD patients significantly increase during the first 3 years. This increment is dependent on or is associated with RRF and PPL (higher levels in patients with lower RRF and higher PPL). FGF-21 might be an important endocrine agent in PD patients and could act as hormonal signaling to maintain glucose homeostasis and prevent potential insulin resistance. These preliminary results suggest that FGF-21 might play a protective role as against the development of insulin resistance over time in patients undergoing a continuous glucose load.     

Liraglutide Increases FGF-21 Activity and Insulin Sensitivity in High Fat Diet and Adiponectin Knockdown Induced Insulin Resistance

Background

Liraglutide is a glucagon-like peptide-1 analogue that stimulates insulin secretion and improves β-cell function. However, it is not clear whether liraglutide achieves its glucose lowering effect only by its known effects or whether other as yet unknown mechanisms are involved. The aim of this study was to examine the effects of liraglutide on Fibroblast growth factor-21 (FGF-21) activity in High-fat diet (HFD) fed ApoE^−/− mice with adiponectin (Acrp30) knockdown.

Method

HFD-fed ApoE^−/− mice were treated with adenovirus vectors expressing shAcrp30 to produce insulin resistance. Hyperinsulinemic-euglycemic clamp studies were performed to evaluate insulin sensitivity of the mouse model. QRT-PCR and Western blot were used to measure the mRNA and protein expression of the target genes.

Results

The combination of HFD, ApoE deficiency, and hypoadiponectinemia resulted in an additive effect on insulin resistance. FGF-21 mRNA expressions in both liver and adipose tissues were significantly increased while FGF-21 receptor 1 (FGFR-1) and β-Klotho mRNA levels in adipose tissue, as well as FGFR-1-3 and β-Klotho mRNA levels in liver were significantly decreased in this model. Liraglutide treatment markedly improved insulin resistance and increased FGF-21 expression in liver and FGFR-3 in adipose tissue, restored β-Klotho mRNA expression in adipose tissue as well as FGFR-1-3, β-Klotho levels and phosphorylation of FGFR1 up to the levels observed in control mice in liver. Liraglutide treatment also further increased FGF-21 proteins in liver and plasma. In addition, as shown by hyperinsulinemic-euglycemic clamp, liraglutide treatment also markedly improved glucose metabolism and insulin sensitivity in these animals.

Conclusion

These findings demonstrate an additive effect of HFD, ApoE deficiency, and adiponectin knockdown on insulin resistance and unveil that the regulation of glucose metabolism and insulin sensitivity by liraglutide may be partly mediated via increased FGF-21 and its receptors action.     

The effect of exercise induced cytokines on insulin stimulated glucose transport in C2C12 cells

Skeletal muscle contractile activity increases the production of the myokines interleukin-6 (IL-6), interleukin-8 (IL-8), interleukin-15 (IL-15) and also skeletal muscle glucose transport. Previous work has revealed a role for IL-6 in mediating glucose uptake, while research on the physiological roles of IL-8 and IL-15 is not so abundant. In the present study we investigated the effects of different concentrations and combinations of IL-6, IL-8 and IL-15 on insulin stimulated glucose transport in C2C12 cells. Furthermore, we also measured AMPK Thr172 and Akt Ser473 phosphorylation via Western blotting. Exposure to 20 pg/ml of individual cytokines had no affect on glucose transport while 1 ng/ml enhanced (P<0.05) glucose uptake with IL-6, IL-8 and IL-15, respectively. Moreover, the combinations of IL-8+IL-6 and IL-15+IL-6 at both 20 pg/ml and 1 ng/ml stimulated (P<0.05) glucose transport with IL-8+IL-15 and IL-8+IL-6+IL-15 only increasing (P<0.05) glucose transport at 1 ng/ml, with no affect observed of these combinations at 20 pg/ml. The changes in glucose transport were all associated with an increase (P<0.05) in AMPK Thr172 phosphorylation with no changes in Akt Ser473 phosphorylation. These findings demonstrated that the exercise induced myokines IL-6, IL-8 and IL-15 enhance glucose transport at 1 ng/ml, with changes only seen at 20 pg/ml with certain myokine combinations. Furthermore these changes in insulin stimulated glucose transport were associated with increased AMPK phosphorylation.     

Subcutaneous adipose tissue exerts proinflammatory cytokines after minimal trauma in humans

Inflammatory cytokines released from adipose tissue play an important role in different pathological processes. In the present study, we investigated the inflammatory cytokine response of human subcutaneous adipose tissue (SAT) by applying the open-flow microperfusion technique. Four standard 18-gauge microperfusion catheters were inserted into periumbilical SAT of eight healthy male volunteers [29 +/- 3 yr, BMI 24.3 +/- 1.9 (mean +/- SD)]. SAT probe effluents were collected at 60-min intervals for 8 h after catheter insertion. Different perfusion fluids were used to measure the local effect of insulin and/or glucose on the cytokine response. SAT probe effluents were analyzed for IL-1beta, IL-6, CXCL8 (IL-8), and TNF-alpha. SAT concentrations of IL-1beta increased 100-fold from 1.0 +/- 0.2 pg/ml (mean +/- SE) to 101.5 +/- 23.2 pg/ml (P < 0.001) after 8 h. A 130-fold increase was observed for CXCL8, from 49 +/- 29 to 6,554 +/- 1,713 pg/ml (P < 0.001). Furthermore, a 20-fold increase of IL-6 was observed within the first 5 h (from 159 +/- 123 to 3,554 +/- 394 pg/ml; P < 0.001), and a significant decline to 2,154 +/- 216 pg/ml (P < 0.01) was seen thereafter. Finally, TNF-alpha increased from 1.4 +/- 0.6 to 2.5 +/- 0.5 pg/ml (P < 0.05) in hour 2 and remained stable thereafter. Local administration of insulin exerted a stimulatory effect on the inflammatory response of IL-6. In conclusion, SAT exerts a highly reproducible and consistent proinflammatory cytokine response after minimally invasive trauma caused by the insertion of a catheter in humans.     

Increased angiotensinogen production in epicardial adipose tissue during cardiac surgery: possible role in a postoperative insulin resistance

Critical illness induces among other events production of proinflammatory cytokines that in turn interfere with insulin signaling cascade and induce insulin resistance on a postreceptor level. Recently, local renin-angiotensin system of adipose tissue has been suggested as a possible contributor to the development of insulin resistance in patients with obesity. The aim of our study was to determine local changes of the renin-angiotensin system of subcutaneous and epicardial adipose tissue during a major cardiac surgery, which may serve as a model of an acute stress potentially affecting endocrine function of adipose tissue. Ten patients undergoing elective cardiac surgery were included into the study. Blood samples and samples of subcutaneous and epicardial adipose tissue were collected at the beginning and at the end of the surgery. Blood glucose, serum insulin and adiponectin levels were measured and mRNA for angiotensinogen, angiotensin-converting enzyme and angiotensin II type 1 receptor were determined in adipose tissue samples using RT PCR. Cardiac surgery significantly increased both insulin and blood glucose levels suggesting the development of insulin resistance, while serum adiponectin levels did not change. Expression of angiotensinogen mRNA significantly increased in epicardial adipose tissue at the end of surgery relative to baseline but remained unchanged in subcutaneous adipose tissue. Fat expression of angiotensin-converting enzyme and type 1 receptor for angiotensin II were not affected by surgery. Our study suggests that increased angiotensinogen production in epicardial adipose tissue may contribute to the development of postoperative insulin resistance.     

Long-term high-fat diet links the regulation of the insulin-sensitizing fibroblast growth factor-21 and visfatin

High-fat diet (HFD) is associated with insulin resistance, hyperinsulinemia, elevated plasma free fatty acid (FFA), and increased risk for atherosclerotic vascular disease. However, the mechanisms underlying the HFD-induced insulin resistance have not been fully clarified. The aim of present study is to evaluate the effects of long-term HFD on the regulation of the insulin-sensitizing fibroblast growth factor-21 (FGF-21) and visfatin in ApoE(-/-) mice. A total of twenty male ApoE(-/-) mice were randomly divided into normal chow diet (NC) or HFD (HF) group for 16 weeks. Euglycemic-hyperinsulinemic clamp was performed to evaluate insulin sensitivity in this animal model. Both mRNA and protein contents of FGF-21 and visfatin were assayed by Quantitative real-time PCR and Western blot. Long-term HFD resulted in the marked abnormality of glucose and lipid metabolism as well as a large decrease in whole-body insulin sensitivity. Accompanied by abnormal glucose-lipid metabolism and aggravated insulin resistance, FGF-21, β-klotho, FGFR1, FGFR3 and FGFR4 mRNA expressions were markedly up-regulated, whereas visfatin mRNA expression was markedly down-regulated in liver and/or adipose tissue of HFD-fed mice. In addition, Western blotting also revealed both up-regulation of the FGF-21 protein and down-regulation of visfatin protein in liver, adipose tissue and plasma of HFD-fed mice. Both FGF-21 and visfatin expression and secretion are regulated by a potent regulator, long-term HFD. And these adipokines are associated with glucose-lipid metabolism and insulin resistance.     

Cytokine regulation of skeletal muscle fatty acid metabolism: effect of interleukin-6 and tumor necrosis factor-alpha

IL-6 and TNF-alpha have been associated with insulin resistance and type 2 diabetes. Furthermore, abnormalities in muscle fatty acid (FA) metabolism are strongly associated with the development of insulin resistance. However, few studies have directly examined the effects of either IL-6 or TNF-alpha on skeletal muscle FA metabolism. Here, we used a pulse-chase technique to determine the effect of IL-6 (50-5,000 pg/ml) and TNF-alpha (50-5,000 pg/ml) on FA metabolism in isolated rat soleus muscle. IL-6 (5,000 pg/ml) increased exogenous and endogenous FA oxidation by approximately 50% (P < 0.05) but had no effect on FA uptake or incorporation of FA into endogenous lipid pools. In contrast, TNF-alpha had no effect on FA oxidation but increased FA incorporation into diacylglycerol (DAG) by 45% (P < 0.05). When both IL-6 (5,000 pg/ml) and insulin (10 mU/ml) were present, IL-6 attenuated insulin's suppressive effect on FA oxidation, increasing exogenous FA oxidation (+37%, P < 0.05). Furthermore, in the presence of insulin, IL-6 reduced the esterification of FA to triacylglycerol by 22% (P < 0.05). When added in combination with IL-6 or leptin (10 microg/ml), the TNF-alpha-induced increase in DAG synthesis was inhibited. In conclusion, the results demonstrate that IL-6 plays an important role in regulating fat metabolism in muscle, increasing rates of FA oxidation, and attenuating insulin's lipogenic effects. In contrast, TNF-alpha had no effect on FA oxidation but increased FA incorporation into DAG, which may be involved in the development of TNF-alpha-induced insulin resistance in skeletal muscle.     

Acute interleukin-6 infusion increases IGFBP-1 but has no short-term effect on IGFBP-3 proteolysis in healthy men

Human conditions of elevated interleukin-6 (IL-6) and transgenic mice overexpressing IL-6 have increased proteolytic degradation of insulin-like growth factor binding protein (IGFBP)-3. In addition, IL-6 alters the hepatic expression of insulin-like growth factor-I (IGF-I) and the IGFBPs in vitro. The aim of the present study was to investigate whether moderately elevated IL-6 levels have short-term effects on circulating IGF-I, IGFBP-1 and IGFBP-3 proteolysis in vivo. Healthy men received a 3-h IL-6 (n = 6) or saline (n = 6) infusion and blood samples were collected prior to and up to 8 h after the start of infusion. Free IGF-I, total IGF-I, IGFBP-1, insulin and cortisol were measured using immunoassays. Serum IGFBP-3 proteolysis was analyzed by Western immunoblot and by in vitro degradation of (125)I-IGFBP-3. We found that IL-6 concentrations reaching approximately 100 pg/ml significantly increased IGFBP-1 after the end of infusion in the absence of changes in insulin. In addition, plasma levels of cortisol were increased in response to IL-6 during and after infusion compared to saline. There was no effect of IL-6 on IGFBP-3 proteolysis, total IGF-I or free dissociable IGF-I. These data suggest that moderately elevated levels of IL-6 such as in the post-operative state or after exercise may contribute to increased levels of IGFBP-1. Although this study does not exclude that high levels and/or prolonged exposure to IL-6 may induce IGFBP-3 proteolysis in sepsis or chronic inflammatory disease, it suggests that IL-6 released from exercising skeletal muscle is not directly involved in proteolysis of circulating IGFBP-3.     

Autocrine modulation of glucose transporter SGLT2 by IL-6 and TNF-α in LLC-PK1 cells

We determined in cultured kidney epithelial cells (LLC-PK(1)) the effects of high glucose, interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α) on mRNA and protein expression of the renal glucose transporters SGLT1 and SGLT2. Cultured monolayers were incubated with similar concentrations of IL-6 and TNF-α to those produced by LLC-PK(1) in the presence of 20 mM glucose. Confluent monolayers with either 5 (controls, C) or 20 mM glucose (high glucose, HG) were incubated in the presence of 5 mM glucose, 20 mM glucose, 10 pg/ml IL-6, or TNF-α alone or in combination. Separate groups with IL-6 and TNF-α were incubated with antibodies to their respective receptors. HG induced an increased SGLT1 mRNA at 48 h (p<0.05 vs. C) and protein expression in 120 h (p<0.05 vs. C). HG also induced an increased SGLT2 mRNA at 72 and 96 h (P<0.05 vs. C) and SGLT2 protein expression at 120 h (p<0.05 vs. C). In C, 10 pg/ml IL-6 or TNF-α did not modify SGLT1 mRNA (n.s vs. in the absence of cytokines). In contrast, cytokines induced an increased expression of SGLT1 protein at 120 h (p<0.05 vs. in the absence of cytokines), and SGLT2 mRNA and protein were increased at 96 and 120 h, respectively (p<0.05 vs. in absence of cytokines). No changes were observed when cells were incubated with cytokines and HG (n.s vs. C). In conclusion, this study showed that SGLT2 increased in the presence of IL-6 and TNF-α, indicating an autocrine modulation of the expression of this transporter by cytokines.     

Influence of TNF-alpha and IL-6 infusions on insulin sensitivity and expression of IL-18 in humans

Inflammation is associated with insulin resistance, and both tumor necrosis factor (TNF)-alpha and interleukin (IL)-6 may affect glucose uptake. TNF induces insulin resistance, whereas the role of IL-6 is controversial. High plasma levels of IL-18 are associated with insulin resistance in epidemiological studies. We investigated the effects of TNF and IL-6 on IL-18 gene expression in skeletal muscle and adipose tissue. Nine human volunteers underwent three consecutive interventions, receiving an infusion of recombinant human (rh)IL-6, rhTNF, and saline. Insulin sensitivity was assessed by measurement of whole body glucose uptake with the stable isotope tracer method during a euglycemic hyperinsulinemic clamp (20 mU.min(-1).kg(-1)), which was initiated 1 h after the IL-6-TNF-saline infusion. Cytokine responses were measured in plasma, muscle, and fat biopsies. Plasma concentrations of TNF and IL-6 increased 10- and 38-fold, respectively, during the cytokine infusions. Whole body insulin-mediated glucose uptake was significantly reduced during TNF infusion but remained unchanged during IL-6 infusion. TNF induced IL-18 gene expression in muscle tissue, but not in adipose tissue, whereas IL-6 infusion had no effect on IL-18 gene expression in either tissue. We conclude that TNF-induced insulin resistance of whole body glucose uptake is associated with increased IL-18 gene expression in muscle tissue, indicating that TNF and IL-18 interact, and both may have important regulatory roles in the pathogenesis of insulin resistance.     

Circulating pro- and anti-inflammatory cytokines in Polish women with gestational diabetes.

In this study we measured serum concentrations of proinflammatory interleukin-6, interleukin-8, and interleukin-18 as well as anti-inflammatory interleukin-10 in 30 pregnant women with normal glucose tolerance, in 32 women with abnormal results of a 50-g glucose challenge test, and in 57 patients with gestational diabetes mellitus. Patients with gestational diabetes had significantly higher IL-6 (median 1.0 [0.7-1.5] vs. 0.7 [0.4-0.8] pg/ml, p=0.001), IL-8 (2.1 [1.1-4.2] pg/ml vs. 0.7 [0.4-0.9] pg/ml, p<0.0001), and IL-18 (249.3 [188.5-318.7] pg/ml vs. 186.7 [139.9-243.9] pg/ml, p=0.005) as well as lower IL-10 levels than healthy pregnant women (0.6 [0.5-1.5] pg/ml vs. 2.9 [1.8-3.2] pg/ml, p<0.0001). After adjusting for glucose, insulin, and BMI values, the differences in IL-8 and IL-18 became insignificant, whereas the differences in IL-6 and IL-10 levels remained highly significant (p<0.0001). The subjects with abnormal glucose challenge test results had higher IL-6 levels (0.9 [0.7-1.3] pg/ml, p=0.005) and similar levels of other cytokines as compared with the women with normal glucose tolerance. Our results suggest an impaired balance between circulating pro- and anti-inflammatory cytokines in patients with gestational diabetes; however, a significant contribution of maternal obesity to the increased levels of IL-8 and IL-18 should be underlined.     

成纤维细胞生长因子21对1型糖尿病动物模型的肝糖代谢影响及机制研究

成纤维细胞生长因子(FGF)-21是FGF家族的成员之一.作为近年发现的一种新的糖代谢调节因子,大量研究表明,FGF-21是一种不依赖胰岛素,能够独立降糖的2型糖尿病治疗潜力型药物.但是,能否应用于1型糖尿病的治疗,国内外目前尚无报道.通过改良传统造模方法,诱导小鼠缓慢产生糖耐量异常,研究FGF-21对此类模型的糖代谢影响及肝糖代谢机制.通过检测FGF-21短期注射和长期注射后模型动物血糖的变化,研究FGF-21在模型动物上对血糖的调控效果.采用实时定量PCR检测FGF-21对模型动物肝脏中葡萄糖转运蛋白(GLUT)1、4 mRNA的表达影响.利用蒽酮法检测模型动物肝脏中糖原合成量.实验结果显示,FGF-21能够调节1型糖尿病动物的血糖水平,并呈剂量依赖性.同时,首次在1型糖尿病动物模型上证实了低剂量FGF-21(0.125 mg/kg)与胰岛素的协同作用效果优于相同剂量FGF-21和胰岛素单独注射的效果.治疗结果表明,FGF-21能够维持1型糖尿病动物模型血糖在正常范围,效果优于胰岛素.实时定量PCR结果发现,与胰岛素上调GLUT4 mRNA表达量不同的是,FGF-21作用动物模型8周后,GLUT1 mRNA表达量显著提高,长期的FGF-21与胰岛素协同注射使GLUT1、4 mRNA表达量同时显著提高.长期FGF-21与胰岛素协同注射组和高剂量FGF-21注射均可显著提高模型动物肝糖原的合成.结果表明,FGF-21促进动物模型糖代谢机制与增加GLUT1表达、增加糖原合成作用有关.为临床应用FGF-21治疗1型糖尿病,增加胰岛素敏感性提供了理论依据.     

成纤维细胞生长因子(FGF)-21改善胰岛素抵抗肝细胞对葡萄糖的吸收和肝糖原的合成

在体外建立胰岛素抵抗肝细胞模型,探讨在胰岛素抵抗状态下成纤维细胞生长因子(FGF)-21对模型细胞糖代谢的影响及机制．将HepG2细胞置于10^-7 mol/L 的胰岛素培养基中培养24 h,建立胰岛素抵抗细胞模型．分别用不同浓度的胰岛素和FGF-21处理模型细胞,采用葡萄糖氧化酶-过氧化物酶(GOD-POD)法检测细胞对葡萄糖的摄取情况,并检查胰岛素与FGF-21的协同作用．利用实时荧光定量PCR检测FGF-21对模型细胞葡萄糖转运蛋白1(GLUT1)mRNA表达的影响,蒽酮法检测模型细胞糖原合成量,探讨FGF-21对胰岛素抵抗细胞模型葡萄糖摄取的影响及机制．结果发现,用高浓度胰岛素处理HepG2细胞24 h后,细胞对胰岛素的敏感性显著降低,说明成功建立了胰岛素抵抗细胞模型,抵抗状态可维持48 h,未发现细胞形态学变化．FGF-21能改善胰岛素抵抗模型细胞的葡萄糖摄取,参与肝糖原的合成,并与胰岛素产生协同作用．实时荧光定量PCR结果发现,FGF-21作用模型细胞后,细胞的GLUT1 mRNA表达量显著增加,说明FGF-21促进模型细胞摄取葡萄糖的作用机制与其增加GLUT1的表达有关．     

Variations of glucose utilization in muscle and adipose tissue of sand rats (Psammomys obesus) during adaptation to laboratory holding

Sand rats, captured in Egypt and fed with a low caloric vegetable diet during adaptation, were investigated before and after 2.5 and 8 weeks diet treatment (30 and 40 kcal/100 g body weight daily). In hexobarbital anaesthesia the sand rats were loaded with 1 g glucose/kg body weight in a single dose intravenously. After a rapid increase the content of glucose in blood remained at a level of about 600 mg glucose/100 ml blood. The insulin immunoreactivity in blood did not change uniformly after application of glucose and remained in a physiologic range. In the islets of Langerhans a degranulation was found during diet treatment. The sensitivity of the epididymal adipose tissue towards insulin in vitro decreased to a nearly complete resistance in the course of diet treatment. A diminution of insulin sensitivity was also found in the m. soleus in vitro. The content of glucose-6-phosphate in the m. semimembranosus was found enhanced after the preparation of the animal. It was found progressively increased up to the five-fold at the end of diet treatment. In the corresponding muscle the glucose distribution volume was increased to about double the extracellular volume. An accumulation of free glucose within the muscle cell must be taken into account. In conclusion the treatment of sand rats with a diabetogenic diet results very quickly in a loss of insulin sensitivity of adipose tissue. The progressively increased stress-mediated accumulation of glucose-6-phosphate and free glucose refers to an inhibition of glucose utilization in the phosphorylation step of glucose in skeletal muscle.     

Plasma inflammatory cytokine response to surgical trauma in chronic depressed patients

We investigated inflammatory cytokine response in chronic depressed patients during abdominal surgery. Twenty-five major depressed patients (Group D) and twenty-five patients (Group C) as the control were studied. Plasma interleukin 6 (IL-6), interleukin 8 (IL-8) and tumour necrosis factor-alpha (TNF-alpha) concentrations were measured before and at 15 min after induction of anesthesia, the end of surgery, 24 h and 3 days after the operation. Plasma IL-6 concentrations in Group D at the end of the operation and 24 h after surgery were significantly lower than those of Group C. The plasma IL-6 concentration (87.1+/-55.3 pg/ml) of patients scoring more than 18 points in the Hamilton depression-rating score at the end of the operation was significantly higher than 57.5+/-76.7 pg/ml of patients scoring less than 18 points. Plasma IL-8 concentration (6.1+/-3.2 pg/ml) in Group D at the end of the operation was significantly lower than 8.7+/-4.2 pg/ml of Group C. We conclude that plasma IL-6 and IL-8 response to surgical trauma is inhibited in chronic depressed patients. The IL-6 response to surgical trauma is depending on the clinical state of depression.     

GDF-15 Is Elevated in Children with Mitochondrial Diseases and Is Induced by Mitochondrial Dysfunction

BackgroundWe previously described increased levels of growth and differentiation factor 15 (GDF-15) in skeletal muscle and serum of patients with mitochondrial diseases. Here we evaluated GDF-15 as a biomarker for mitochondrial diseases affecting children and compared it to fibroblast-growth factor 21 (FGF-21). To investigate the mechanism of GDF-15 induction in these pathologies we measured its expression and secretion in response to mitochondrial dysfunction.MethodsWe analysed 59 serum samples from 48 children with mitochondrial disease, 19 samples from children with other neuromuscular diseases and 33 samples from aged-matched healthy children. GDF-15 and FGF-21 circulating levels were determined by ELISA.ResultsOur results showed that in children with mitochondrial diseases GDF-15 levels were on average increased by 11-fold (mean 4046pg/ml, 1492 SEM) relative to healthy (350, 21) and myopathic (350, 32) controls. The area under the curve for the receiver-operating-characteristic curve for GDF-15 was 0.82 indicating that it has a good discriminatory power. The overall sensitivity and specificity of GDF-15 for a cut-off value of 550pg/mL was 67.8% (54.4%-79.4%) and 92.3% (81.5%-97.9%), respectively. We found that elevated levels of GDF-15 and or FGF-21 correctly identified a larger proportion of patients than elevated levels of GDF-15 or FGF-21 alone. GDF-15, as well as FGF-21, mRNA expression and protein secretion, were significantly induced after treatment of myotubes with oligomycin and that levels of expression of both factors significantly correlated.ConclusionsOur data indicate that GDF-15 is a valuable serum quantitative biomarker for the diagnosis of mitochondrial diseases in children and that measurement of both GDF-15 and FGF-21 improves the disease detection ability of either factor separately. Finally, we demonstrate for the first time that GDF-15 is produced by skeletal muscle cells in response to mitochondrial dysfunction and that its levels correlate in vitro with FGF-21 levels.     

Suppressing lipolysis increases interleukin-6 at rest and during prolonged moderate-intensity exercise in humans.

IL-6 induces lipolysis when administered to humans. Consequently, it has been hypothesized that IL-6 is released from skeletal muscle during exercise to act in a "hormonelike" manner and increase lipolysis from adipose tissue to supply the muscle with substrate. In the present study, we hypothesized that suppressing lipolysis, and subsequent free fatty acid (FFA) availability, would result in a compensatory elevation in IL-6 at rest and during exercise. First, we had five healthy men ingest nicotinic acid (NA) at 30-min intervals for 120 min at rest [10 mg/kg body mass (initial dose), 5 mg/kg body mass (subsequent doses)]. Plasma was collected and analyzed for FFA and IL-6. After 120 min, plasma FFA concentration was attenuated (0 min: 0.26 +/- 0.05 mmol/l; 120 min: 0.09 +/- 0.02 mmol/l; P < 0.01), whereas plasma IL-6 was concomitantly increased approximately eightfold (0 min: 0.75 +/- 0.18 pg/ml; 120 min: 6.05 +/- 0.89 pg/ml; P < 0.001). To assess the effect of lipolytic suppression on the exercise-induced IL-6 response, seven active, but not specifically trained, men performed two experimental exercise trials with (NA) or without [control (Con)] NA ingestion 60 min before (10 mg/kg body mass) and throughout (5 mg/kg body mass every 30 min) exercise. Blood samples were obtained before ingestion, 60 min after ingestion, and throughout 180 min of cycling exercise at 62 +/- 5% of maximal oxygen consumption. IL-6 gene expression, in muscle and adipose tissue sampled at 0, 90, and 180 min, was determined by using semiquantitative real-time PCR. IL-6 mRNA increased in Con (rest vs. 180 min; P < 0.01) approximately 13-fold in muscle and approximately 42-fold in fat with exercise. NA increased (rest vs. 180 min; P < 0.01) IL-6 mRNA 34-fold in muscle, but the treatment effect was not statistically significant (Con vs. NA, P = 0.1), and 235-fold in fat (Con vs. NA, P < 0.01). Consistent with the study at rest, NA completely suppressed plasma FFA (180 min: Con, 1.42 +/- 0.07 mmol/l; NA, 0.10 +/- 0.01 mmol/l; P < 0.001) and increased plasma IL-6 (180 min: Con, 9.81 +/- 0.98 pg/ml; NA, 19.23 +/- 2.50 pg/ml; P < 0.05) during exercise. In conclusion, these data demonstrate that circulating IL-6 is markedly elevated at rest and during prolonged moderate-intensity exercise when lipolysis is suppressed.     

Plasma proinflammatory cytokine response to surgical stress in elderly patients

We investigated the change of plasma cytokines concentrations in elderly patients during lower abdominal surgery. Plasma interleukin (IL-)6 concentrations (68.0+/-15.4 and 36.1+/-20.7 pg/ml) in elderly patients at 24 h and at 3 days after surgery were significantly higher than those (35.1+/-21.5 and 18.6+/-10.6 pg/ml) of young adults. Plasma IL-6 concentrations (92.3+/- 31.9 pg/ml) in elderly patients anesthetized with propofol and fentanyl were significantly higher at the end of the operation than that (57.9+/-36.7 pg/ml) of elderly patients anesthetized with sevoflurane and fentanyl. In conclusion, elderly patients have an increased and delayed IL-6 response to surgical trauma compared with young adults. Plasma IL-6 production after surgical trauma in elderly patients with total intravenous anesthesia with propofol was significantly higher than that in elderly patients with sevoflurane anesthesia.