The Growth Hormone Releasing Hormone Analogue, Tesamorelin, Decreases Muscle Fat and Increases Muscle Area in Adults with HIV.

BACKGROUND: Tesamorelin, a growth hormone-releasing hormone analogue, decreases visceral adipose tissue in people living with HIV, however, the effects on skeletal muscle fat and area are unknown. OBJECTIVES: The goals of this exploratory secondary analysis were to determine the effects of tesamorelin on muscle quality (density) and quantity (area). DESIGN: Secondary, exploratory analysis of two previously completed randomized (2:1), clinical trials. SETTING: U.S. and Canadian sites. PARTICIPANTS: People living with HIV and with abdominal obesity. Tesamorelin participants were restricted to responders (visceral adipose tissue decrease ≥8%). INTERVENTION: Tesamorelin or placebo. MEASUREMENTS: Computed tomography scans (at L4-L5) were used to quantify total and lean density (Hounsfield Units, HU) and area (centimeters2) of four trunk muscle groups using a semi-automatic segmentation image analysis program. Differences between muscle area and density before and after 26 weeks of tesamorelin or placebo treatment were compared and linear regression models were adjusted for baseline and treatment arm. RESULTS: Tesamorelin responders (n=193) and placebo (n=148) participants with available images were similar at baseline; most were Caucasian (83%) and male (87%). In models adjusted for baseline differences and treatment arm, tesamorelin was associated with significantly greater increases in density of four truncal muscle groups (coefficient 1.56-4.86 Hounsfield units; all p<0.005), and the lean anterolateral/abdominal and rectus muscles (1.39 and 1.78 Hounsfield units; both p<0.005) compared to placebo. Significant increases were also seen in total area of the rectus and psoas muscles (0.44 and 0.46 centimeters2; p<0.005), and in the lean muscle area of all four truncal muscle groups (0.64-1.08 centimeters2; p<0.005). CONCLUSIONS: Among those with clinically significant decrease in visceral adipose tissue on treatment, tesamorelin was effective in increasing skeletal muscle area and density. Long term effectiveness of tesamorelin among people with and without HIV, and the impact of these changes in daily life should be further studied.

Advanced glycation end-products increase monocyte adhesion to retinal endothelial cells through vascular endothelial growth factor-induced ICAM-1 expression: inhibitory effect of antioxidants.

Accumulating evidence indicates a role for advanced glycation end-products (AGEs) in the development of diabetic retinopathy. In the present study, we examined the in vitro effect of AGEs on human monocyte adhesion to bovine retinal endothelial cells (BRECs) and the molecular mechanisms involved in this effect. Treatment of cultured BRECs with AGEs led to a significant increase in monocyte adhesion and intercellular cell adhesion molecule-1 (ICAM-1) expression. These effects were inhibited by antioxidants including gliclazide and vitamins C and E. On the basis of the stimulatory effect of AGEs on vascular endothelial growth factor (VEGF) secretion by retinal endothelial cells, the role of this growth factor as mediator of AGE-induced monocyte adhesion to BRECs was next investigated. Incubation of BRECs with VEGF increased monocyte adhesion to these cells and enhanced ICAM-1 expression. Treatment of BRECs with an anti-VEGF antibody abrogated AGE-induced monocyte adhesion and ICAM-1 expression. Finally, incubation of BRECs with protein kinase C (PKC) and nuclear factor (NF)-kappaB inhibitors suppressed monocyte adhesion and ICAM-1 expression elicited by AGEs and VEGF. Taken together, these data indicate that AGEs increase monocyte adhesion to BRECs and that this effect is mediated through VEGF-induced ICAM-1 expression. They also demonstrate that this effect is oxidative stress-sensitive and involves PKC and NF-kappaB-dependent signaling pathways.

Signalling pathways involved in retinal endothelial cell proliferation induced by advanced glycation end products: inhibitory effect of gliclazide.

AIM: We have previously demonstrated that advanced glycation end products (AGEs) stimulate bovine retinal endothelial cell (BREC) proliferation through induction of vascular endothelial growth factor (VEGF) production by these cells. We have also shown that gliclazide, a sulfonylurea which decreases oxidative stress, inhibits this effect. The aim of the present study was to characterize the signalling pathways involved in AGE-induced BREC proliferation and VEGF production and mediating the inhibitory effect of gliclazide on these biological events. METHODS: BRECs were treated or not treated with AGEs in the presence or absence of gliclazide, antioxidants, protein kinase C (PKC), mitogen-activated protein kinase (MAPK) or nuclear factor-kappaB (NF-kappaB) inhibitors. BREC proliferation was assessed by measuring [3H]-thymidine incorporation into DNA. Activation of PKC, MAPK and NF-kappaB signal transduction pathways and determination of VEGF expression were assessed by Western blot analysis using specific antibodies. MAPK activity was also determined by an in vitro kinase assay. RESULTS: Treatment of BRECs with AGEs significantly increased cell proliferation and VEGF expression. AGEs induced PKC-beta translocation, extracellular signal-regulated protein kinase 1/2 and NF-kappaB activation in these cells. Pharmacological inhibition of these signalling pathways abolished AGE effects on cell proliferation and VEGF expression. Exposure of BRECs to gliclazide or antioxidants such as vitamin E or N-acetyl-l-cysteine resulted in a significant decrease in AGE-induced activation of PKC-, MAPK- and NF-kappaB-signalling pathways. CONCLUSIONS: Our results demonstrate the involvement of PKC, MAPK and NF-kappaB in AGE-induced BREC proliferation and VEGF expression. Gliclazide inhibits BREC proliferation by interfering with these intracellular signal transduction pathways.

Lack of concordance between plasma markers of cardiovascular risk and intima-media thickness in patients with type 2 diabetes.

AIM: Endothelial dysfunction, oxidative stress and systemic inflammation play an important role in the enhanced cardiovascular risk in diabetes. Carotid intima-media thickness (IMT), a widely accepted marker of subclinical atherosclerosis, is known to be increased in patients with type 2 diabetes. The relationships between plasma markers of cardiac risk and carotid IMT are not well known. We therefore studied a group of patients with type 2 diabetes to evaluate the relationships between plasma markers of cardiac risk and carotid IMT. DESIGN AND PATIENTS: We measured carotid IMT and the levels of soluble endothelial adhesion molecules [sE-selectin, intercellular cell adhesion molecule-1 (sICAM-1) and vascular cell adhesion molecule-1 (sVCAM-1)], C-reactive protein (CRP) and 8-isoprostane in 40 patients with type 2 diabetes without clinical macrovascular complications (HbA1c<10%, duration of diabetes<12 years) and 25 healthy subjects. We then examined the correlations between these plasma markers, carotid IMT and various clinical and biochemical parameters. RESULTS: Diabetic patients had higher plasma sE-selectin (p=0.03), sICAM-1 (p=0.05), CRP (p=0.047) and 8-isoprostane (p=0.001) concentrations than control subjects. Mean IMT values were identical (0.63 +/- 0.02 mm) in diabetic (range, 0.40-0.92 mm) and healthy subjects (range, 0.45-0.85 mm). In diabetic patients, stepwise multivariate analysis showed that HbA1c and plasma glucose were independent predictors of sE-selectin (r2=0.19 and r2=0.17, p<0.01, respectively), whereas waist circumference and body mass index (BMI) were predictors of sICAM-1 (r2=0.27, p=0.001 and r2=0.22, p=0.002, respectively). Waist circumference was the only predictor of CRP (r2=0.2, p<0.01), and systolic blood pressure was the only predictor of 8-isoprostane (r2=0.19, p=0.006). In control subjects, similar analysis showed that plasma glucose and waist circumference were predictors of sE-selectin and sICAM-1, respectively (r2=0.2, p<0.05). CONCLUSIONS: These results indicate that some well-controlled type 2 diabetic patients free of clinical macrovascular complications have elevated plasma markers of cardiovascular risk without having increased IMT. The elevation of plasma markers of endothelial cell activation (sE-selectin and s-ICAM-1) or inflammation (CRP) and oxidative stress (8-isoprostane) in diabetics vs. controls is distinct from and cannot be explained simply by differences in the burden of atherosclerosis as assessed by carotid IMT.

Antiatherogenic properties of metformin: the experimental evidence.

Cardiovascular disease (CVD) is the major determining factor of morbidity and mortality in type 2 diabetic patients. The established relationship between type 2 diabetes and atherosclerosis has fueled suggestions that anti-diabetic drugs with beneficial effects on CV risk factors may help attenuate the atherosclerotic process in diabetic patients. Metformin is a hypoglycaemic agent widely used in the management of type 2 diabetes. In addition to its insulin-sensitising action, this drug has favourable effects on various CV risk factors and reduces macrovascular complications in obese type 2 diabetic patients. This review summarises in vivo and in vitro experimental evidence on the antiatherogenic properties of metformin.

Gliclazide decreases vascular smooth muscle cell dysfunction induced by cell-mediated oxidized low-density lipoprotein.

Accumulating evidence indicates that oxidative modification of low-density lipoprotein (LDL) plays an important role in vascular dysfunction associated with diabetes mellitus. The aim of the present study was to investigate the effect of gliclazide, a second-generation sulfonylurea with free-radical-scavenging activity, on human aortic smooth muscle cell (HASMC)-mediated LDL oxidation and HASMC dysfunction induced by oxidatively modified LDL. Incubation of HASMCs with native human LDL (100 microg/mL) in the presence of increasing concentrations of gliclazide (1 to 10 microg/mL) resulted in a dose-dependent decrease in HASMC-mediated LDL oxidation. Exposure of HASMCs to gliclazide (1 to 10 microg/mL) and native LDL (100 microg/mL) also led to a dose-dependent decrease in oxidized LDL-induced human monocyte adhesion to HASMCs. In addition, incubation of HASMCs with gliclazide dramatically reduced the ability of oxidized LDL to stimulate the proliferation of these cells. Finally, treatment of HASMCs with gliclazide resulted in a marked decrease in oxidatively modified LDL-induced monocyte chemoattractant protein (MCP)-1 and human heat shock protein 70 (hsp 70) expression, both at the gene and protein levels. These results show that gliclazide, at concentrations in the therapeutic range (5 to 10 microg/mL), is effective in vitro in reducing vascular smooth muscle cell (VSMC) dysfunction induced by oxidatively modified LDL. These observations suggest that administration of gliclazide to type 2 diabetic patients could form part of the strategy for the prevention and management of diabetic cardiovascular diseases.

Proliferative effect of lipoprotein lipase on human vascular smooth muscle cells.

Vascular smooth muscle cell (VSMC) proliferation is a key event in the development and progression of atherosclerotic lesions. Accumulating evidence suggests that lipoprotein lipase (LPL) produced in the vascular wall may exert proatherogenic effects. The aim of the present study was to examine the effect of LPL on VSMC proliferation. Incubation of growth-arrested human VSMCs with purified endotoxin-free bovine LPL for 48 and 72 hours, in the absence of any added exogenous lipoproteins, resulted in a dose-dependent increase in VSMC growth. Addition of VLDLs to the culture media did not further enhance the LPL effect. Treatment of growth-arrested VSMCs with purified human or murine LPL (1 microg/mL) led to a similar increase in cell proliferation. Neutralization of bovine LPL by the monoclonal 5D2 antibody, irreversible inhibition, or heat inactivation of the lipase suppressed the LPL stimulatory effect on VSMC growth. Moreover, preincubation of VSMCs with the specific protein kinase C inhibitors calphostin C and chelerythrine totally abolished LPL-induced VSMC proliferation. In LPL-treated VSMCs, a significant increase in protein kinase C activity was observed. Treatment of VSMCs with heparinase III (1 U/mL) totally inhibited LPL-induced human VSMC proliferation. Taken together, these data indicate that LPL stimulates VSMC proliferation. LPL enzymatic activity, protein kinase C activation, and LPL binding to heparan sulfate proteoglycans expressed on VSMC surfaces are required for this effect. The stimulatory effect of LPL on VSMC proliferation may represent an additional mechanism through which the enzyme contributes to the progression of atherosclerosis.

Differentiation of human monocytes to monocyte-derived macrophages is associated with increased lipoprotein lipase-induced tumor necrosis factor-alpha expression and production: a process involving cell surface proteoglycans and protein kinase C.

The aim of the present study was to (1) evaluate the responsiveness of human mononuclear cells to lipoprotein lipase (LPL), as assessed by tumor necrosis factor-alpha (TNFalpha) production, during the process of differentiation of monocytes to macrophages, and (2) determine the mechanisms by which LPL exerts its effect on these cells. Treatment of human monocytes with purified endotoxin-free bovine LPL (1 microgram/mL) resulted in a 161+/-15% increase in TNFalpha production over control values (P<0.01). A further increase in TNFalpha production was observed after treatment of monocyte-derived macrophages (MDMs) with LPL (490+/-81% over control values, P<0.01). Increased TNFalpha mRNA expression and protein kinase C activity were also observed in LPL-treated human monocytes and MDMs. These LPL effects were abrogated by the specific protein kinase C inhibitor calphostin C (1 micromol/L). Although heparinase totally abolished LPL-induced TNFalpha production in human monocytes, this agent did not significantly inhibit LPL effect in human MDMs. In contrast, treatment of MDMs with chondroitinase suppressed LPL-induced TNFalpha production. Taken together, these data suggest that (1) differentiation of human monocytes to MDMs is associated with increased LPL-induced TNFalpha mRNA expression and production, (2) a protein kinase C-dependent pathway is involved in the induction of TNFalpha by LPL in these cells, and (3) LPL effect is mediated by cell surface proteoglycans. As MDMs secrete LPL in the vascular wall, we propose that LPL, by acting as an autocrine activator of MDM function, may contribute to the high level of TNFalpha found in the atheromatous lesion.

Lipoprotein lipase enhances human monocyte adhesion to aortic endothelial cells.

Lipoprotein lipase (LPL)-mediated lipolysis of very low density lipoprotein (VLDL) has been demonstrated to increase U937 monocyte adhesion to endothelial cells. In the present study, we evaluated the ability of LPL to enhance human monocyte adhesion to bovine aortic endothelial cells (BAEC) in the absence of exogenous lipoproteins. Exposure of BAEC to 1 microgram/ml LPL at 37 degrees C resulted in a significant increase in monocyte adhesion over control values. Addition of VLDL in the culture media further enhanced the LPL effect. A significant increase in monocyte adhesion was also observed when BAEC were incubated with LPL at 4 degrees C. Heparin or heparinase treatment of BAEC totally abolished the LPL stimulatory effect on monocyte adhesion. In addition, incubation of monocytes with heparinase suppressed the ability of LPL to stimulate monocyte adhesion to endothelial cells. These treatments also markedly decreased LPL binding to the monocyte and endothelial cell surfaces. In contrast to native LPL, heat inactivated or phenylmethylsulfonyl fluoride (PMSF)-treated LPL did not increase monocyte adhesion to BAEC. Finally, incubation of LPL in the presence of the 5D2 antibody resulted in a total suppression of the LPL-induced monocyte adhesion to BAEC. Taken together, these data demonstrate that LPL activity plays an important role in LPL-induced monocyte adhesion and that LPL binding to heparan sulfate proteoglycans expressed on both monocytes and endothelial cells surfaces is required for the enhanced monocyte adhesion. These results suggest a new mechanism by which LPL may promote the development of atherosclerosis, that of facilitating monocyte adhesion to the endothelium.