Thiazolidinedione Treatment Research Articles

Mechanisms of adipose tissue redistribution with rosiglitazone treatment in various adipose depots

Treatment with thiazolidinediones (TZDs) improves glucose homeostasis by increasing insulin sensitivity, but it also leads to weight gain. Our hypothesis was that, in individual adipose depots, there is depot specificity for lipid storage and energy expenditure genes after TZD treatment. After 5 weeks of rosiglitazone treatment on Otsuka Long-Evans Tokushima Fatty (OLETF) rats, an animal model of type 2 diabetes mellitus with obesity, and Long-Evans Tokushima Otsuka rats as controls, we measured changes in lipid storage and energy expenditure gene expression in various adipose depots, such as mesenteric and nonmesenteric adipose tissues (subcutaneous, epididymal, and retroperitoneal). Mesenteric fat masses did not change after TZD treatment in OLETF rats, but nonmesenteric fat masses increased. Messenger RNA expression of lipid storage genes increased in nonmesenteric fat, but energy expenditure gene expression increased in mesenteric fat after rosiglitazone treatment. In conclusion, our findings suggest that TZD treatment may be associated with the depot-specific effects of lipid storage and energy expenditure genes on fat redistribution in individual adipose tissues in OLETF rats.

Down-regulation of lipoprotein lipase increases glucose uptake in L6 muscle cells

Thiazolidinediones (TZDs) are synthetic hypoglycemic agents used to treat type 2 diabetes. TZDs target the peroxisome proliferator activated receptor-gamma (PPAR-γ) and improve systemic insulin sensitivity. The contributions of specific tissues to TZD action, or the downstream effects of PPAR-γ activation, are not very clear. We have used a rat skeletal muscle cell line (L6 cells) to demonstrate that TZDs directly target PPAR-γ in muscle cells. TZD treatment resulted in a significant repression of lipoprotein lipase (LPL) expression in L6 cells. This repression correlated with an increase in glucose uptake. Down-regulation of LPL message and protein levels using siRNA resulted in a similar increase in insulin-dependent glucose uptake. Thus, LPL down-regulation improved insulin sensitivity independent of TZDs. This finding provides a novel method for the management of insulin resistance.

Bone Disease, Gestational Diabetes Mellitus, and Health Care

This is the seventh of a series of articles based on presentations at the American Diabetes Association (ADA) Scientific Sessions held 6–10 June 2008 in San Francisco, California. ### Type 2 diabetes and bone disease Robert Josse (Toronto, Canada) discussed new concepts of skeletal homeostasis and its disorders in a symposium addressing the relationship of type 2 diabetes, and in particular that of thiazolidinedione (TZD) treatment, to bone disease. Remodeling is the process of combined osteoclast and osteoblast activity that optimizes bone structure to improve strength and mechanical function and repair microdamage, fulfills metabolic functions, and acts as an important source of growth factors. The basic multicellular units of bone contain osteoclasts that excavate bone, mononuclear cells that remove cellular debris, and osteoblasts that replace the removed bone. Receptor activator of nuclear factor-κB (RANK) ligand is a transmembrane protein belonging to the tumor necrosis factor superfamily that specifically binds RANK and osteoprotegerin and plays an important role in regulating osteoclast differentiation and activation, whereas the Wnt (related to a gene controlling wing forming in fruit flies) system controls osteoblast activity with complex signals linking osteoblasts and osteoclasts (1). An LDL receptor–related protein (LRP)5 mutation is associated with increased bone mass. The LRP5/Wnt signaling pathway increases activity of the multifunctional protein β-catenin that participates in cell adhesion and nuclear signaling and is inhibited by an osteocyte product sclerostin (defects in this are associated with sclerositis) and by the antagonist Dickkopf (DKK) proteins. DKK1 is a myeloblast product that helps explain the failure of multiple myeloma lesions to show activity on bone scan. Osteoporosis may be caused by increased osteoclast or decreased osteoblast activity, compromising bone strength, which Josse suggested be seen as an integrated measure of bone density and bone quality, recognizing that standard bone density measurement fails to assess the latter component. Bones break when the applied …

Tumor Necrosis Factor-α Induces Caspase-mediated Cleavage of Peroxisome Proliferator-activated Receptor γ in Adipocytes

The nuclear receptor peroxisome proliferator-activated receptor gamma (PPARgamma) is a ligand-dependent transcription factor that acts as a primary regulator of adipogenesis and controls adipocyte metabolism and insulin action. Increased expression of tumor necrosis factor (TNFalpha) in adipose tissue of obese subjects potently suppresses the expression of PPARgamma and attenuates adipocyte functions. Here we show that PPARgamma is a substrate of caspase-3 and caspase-6 during TNFalpha receptor signaling in adipocytes, and the consequent PPARgamma cleavage disrupts its nuclear localization. TNFalpha treatment of 3T3-L1 adipocytes decreases full-length PPARgamma while increasing the level of a 45-kDa immunoreactive PPARgamma fragment. Specific inhibitors of caspase-3 and caspase-6 attenuate the cleavage of PPARgamma protein in response to TNFalpha in cultured adipocytes. Incubation of nuclear fractions with recombinant caspase-3 and caspase-6 also generates a 45-kDa PPARgamma cleavage product. Dispersion of nuclear PPARgamma to the cytoplasm in response to TNFalpha treatment occurs in parallel with detection of activated caspase-3. We suggest that activation of the caspase cascade by TNFalpha down-regulates PPARgamma protein and PPARgamma-mediated metabolic processes in adipose cells.

Thiazolidinediones may contribute to the intramyocardial lipid accumulation in diabetic myocardium: effects on cardiac function

To the editor: Treatment with thiazolidinedione (TZD), an agonist of peroxisome proliferator-activated receptor-γ (PPARγ), has been associated with a significant increase in heart failure in type 2 diabetic patients.1 These...

Thiazolidinedione use is associated with better survival in hemodialysis patients with non-insulin dependent diabetes

Cardiovascular mortality is especially high among dialysis patients with diabetes, as is morbidity due to protein energy wasting. Given that both of these factors may be decreased by thiazolidinedione treatment, we studied the effect of thiazolidinedione use on survival among chronic dialysis patients in a national cohort of 5290 incident dialysis patients with diabetes. Thiazolidinedione use was assessed according to prescription data, and the analyses were stratified based on insulin use due to observed interaction. In the primary analysis, thiazolidinedione treatment was associated with significantly lower all-cause mortality among insulin-free but not insulin-requiring subjects, with adjusted hazards ratios of 0.53 (0.31-0.89) and 0.82 (0.46-1.47) respectively. Sensitivity analyses found the findings to be robust with respect to confounding by indication, severity of the diabetes, potential reverse causality, and time varying exposure patterns. The mechanism of this decline in all-cause mortality will need to be examined after these studies are confirmed.

Altering PPARγ Ligand Selectivity Impairs Adipogenesis by Thiazolidinediones But Not Hormonal Inducers

Peroxisome proliferator-activated receptor gamma (PPARgamma) acts as a ligand-dependent transcription factor with a key role in mediating adipocyte differentiation and insulin sensitivity. Recently, we and others have shown that PPARgamma recruits the nuclear corepressors NCoR and silencing mediator for retinoid and thyroid hormone receptors (SMRT) to modulate adipogenesis. While the synthetic ligands for PPARgamma, the thiazolidinediones (TZD), are widely used in the treatment of type 2 diabetes mellitus, the biologically relevant endogenous PPARgamma ligand involved in adipogenesis remains unidentified. To further understand the role of ligand binding and corepressor interaction in PPARgamma-mediated adipogenesis, a mutation was introduced in the ligand-binding domain (LBD) of murine PPARgamma. PPARgammamut was created via two amino acid substitutions known to be major determinants of ligand selectivity among PPAR isotypes, H323Y and R288M. These mutations alter PPARgamma to the corresponding residues of the PPARalpha. Characterizing the in vitro functional properties of this mutant, we show that PPARgammamut preferentially responds to the PPARalpha agonist, WY-14643, over the TZD, pioglitazone. When expressed in 3T3-L1 preadipocytes using recombinant adenovirus, wild-type PPARgamma leads to adipocyte formation with both hormonal and TZD treatment. PPARgammamut blocks the upregulation of adipocyte-specific proteins by TZD, but surprisingly, not by standard hormonal inducers. Our data suggest that TZDs and the purported endogenous ligand do not interact in the same way with the PPARgamma LBD. We propose that the endogenous ligand has distinct properties that allow for promiscuity within the hydrophobic PPAR ligand-binding pocket, yet fosters appropriate cofactor recruitment and release to allow adipogenesis to proceed.

Translating science into clinical practice: focus on vildagliptin in combination with metformin

Vildagliptin is a potent and selective oral dipeptidyl peptidase-4 inhibitor that improves glycaemic control in patients with type 2 diabetes mellitus (T2DM) by increasing both alpha- and beta-cell responsiveness to glucose. The efficacy, tolerability and safety of the combination of vildagliptin and metformin in the treatment of T2DM have been established in numerous trials in the extensive vildagliptin clinical programme. As add-on therapy in patients with inadequate glycaemic control on metformin, vildagliptin produces clinically significant reductions in glycated haemoglobin (HbA1c) and fasting plasma glucose, is well tolerated, and is associated with absence of weight gain and minimal risk of hypoglycaemia. Compared with thiazolidinedione add-on treatment, vildagliptin is associated with similar significant reductions in HbA1c without the weight gain seen with the former. Compared with sulfonylurea add-on treatment, vildagliptin is associated with similar efficacy in controlling glycaemia but absence of weight gain and a markedly lower risk of hypoglycaemia. In drug-naive patients, single-tablet combinations of vildagliptin/metformin 50/500 and 50/1000 mg bid produced significantly greater reductions in HbA1c than monotherapy with either agent and were well tolerated, with no weight gain and minimal risk of hypoglycaemia. The combination of vildagliptin and metformin poses numerous advantages in the treatment of T2DM.

Retinol saturase promotes adipogenesis and is downregulated in obesity

Adipocyte differentiation is controlled by many transcription factors, but few known downstream targets of these factors are necessary for adipogenesis. Here we report that retinol saturase (RetSat), which is an enzyme implicated in the generation of dihydroretinoid metabolites, is induced during adipogenesis and is directly regulated by the transcription factor peroxisome proliferator activated receptor gamma (PPARgamma). Ablation of RetSat dramatically inhibited adipogenesis but, surprisingly, this block was not overcome by the putative product of RetSat enzymatic activity. On the other hand, ectopic RetSat with an intact, but not a mutated, FAD/NAD dinucleotide-binding motif increased endogenous PPARgamma transcriptional activity and promoted adipogenesis. Indeed, RetSat was not required for adipogenesis when cells were provided with exogenous PPARgamma ligands. In adipose tissue, RetSat is expressed in adipocytes but is unexpectedly downregulated in obesity, most likely owing to infiltration of macrophages that we demonstrate to repress RetSat expression. Thiazolidinedione treatment reversed low RetSat expression in adipose tissue of obese mice. Thus, RetSat plays an important role in the biology of adipocytes, where it favors normal differentiation, yet is reduced in the obese state. RetSat is thus a novel target for therapeutic intervention in metabolic disease.

Retinoid and thiazolidinedione therapies in melanoma: an analysis of differential response based on nuclear hormone receptor expression

BackgroundMetastatic melanoma has a high mortality rate and suboptimal therapeutic options. Molecular targeting may be beneficial using the rexinoid LGD1069, a retinoid × receptor selective agonist, and thiazolidinediones (TZD), PPARγ selective ligands, as novel treatments.ResultsMouse xenograft models with human melanoma cell lines [A375(DRO) or M14(5–16)] were treated for 4 weeks with daily vehicle, RXR agonist (rexinoid, LGD1069, 30 mg/kg/d), PPARγ agonist (TZD, rosiglitazone, 10 mg/kg/d) or combination. A375(DRO) tumor growth was significantly inhibited by either ligand alone and the combination had an additive effect. M14(5–16) tumors only responded to LGD1069 100 mg/kg/day. A375(DRO) sublines resistant to rexinoid, TZD and combination were generated and all three sublines had reduced PPARγ expression but preserved RXR expression. shRNA knockdown of PPARγ or RXRγ attenuated the rexinoid, TZD and combination ligand-mediated decreased proliferation in A375(DRO) cells. Rexinoid (LGD1069) and retinoid (TTNPB) treatment of M14(5–16) cells resulted in decreased proliferation that was additive with combination of both rexinoid and retinoid. shRNA knockdown of RXRγ resulted in a decreased response to either ligand.ConclusionA375 (DRO) melanoma cell growth is inhibited by rexinoid and TZD treatment, and this response is dependent on RXR and PPARγ receptor expression. M14 (5–16) melanoma cell growth is inhibited by rexinoid and retinoid treatment, and this response is dependent on RXR expression. These findings may help guide molecular-based treatment strategies in melanoma and provide insight for mechanisms of resistance to nuclear receptor targeted therapies in certain cancers.

Herzinsuffizienz unter Glitazonen – was wissen wir heute?

Patients with type 2 diabetes are at a high risk for cardiovascular disease. A common co-morbidity is heart failure, resulting in increased mortality rates. Research on thiazolidinediones (glitazones) has revealed protective effects on the cardiovascular system. On the other hand, this antidiabetic class causes fluid retention that may promote congestive heart failure. The clinical manifestation of heart failure occurs more frequently on either pioglitazone or rosiglitazone treatment than with other oral antidiabetic drugs. This article provides an overview of the pathophysiological background of fluid retention, the incidence of congestive heart failure with thiazolidinedione treatment and the clinical outcomes in type 2 diabetes with and without a pre-existing condition. A review of the currently available data discloses the following relevant facts: treatment of diabetes with a thiazolidinedione appears to increase the signs of congestive heart failure, but not the risk of cardiovascular and overall death. On the contrary, type 2 diabetics with pre-existing heart failure benefits from a reduction in cardiovascular end-points. All these data suggest that heart failure precipitated by thiazolidinedione drugs is qualitatively different from other causes. This implies the need for a new prognostic evaluation of patients with thiazolidinedione-promoted heart failure.

A pilot study of the effects of pioglitazone and rosiglitazone on de novo lipogenesis in type 2 diabetes

Treatment of type 2 diabetes mellitus (T2DM) patients with pioglitazone results in a more favorable lipid profile, and perhaps more favorable cardiac outcomes, than treatment with rosiglitazone. Pioglitazone treatment increases VLDL-triacylglycerol clearance, but the role of de novo lipogenesis (DNL) has not been explored, and no direct comparison has been made between the thiazolidinediones (TZDs). Twelve subjects with T2DM and hypertriacylglyceridemia were randomized to either rosiglitazone or pioglitazone treatment. Stable isotope infusion studies were performed at baseline and after 20 weeks of treatment. Both treatments reduced glucose and HbA(1c) concentrations equally. Pioglitazone treatment resulted in a 40% reduction in hepatic DNL (P < 0.01) and in a 25% reduction in hepatic glucose production (P < 0.05), while rosiglitazone did not significantly change either parameter, although comparisons of changes between treatments were not significantly different. These pilot results indicate that pioglitazone reduces hepatic DNL while rosiglitazone does not. Larger follow-up studies are required to confirm differential effects of these agents definitively. The reduction in DNL may underlie altered assembly or atherogenicity of lipoprotein particles and may reflect PPARalpha or other non-PPARgamma actions on the liver by pioglitazone. These differences might help explain previously reported differences in lipid profiles and cardiovascular disease outcomes for rosiglitazone and pioglitazone.

Vildagliptin: a new oral treatment for type 2 diabetes mellitus

Vildagliptin is a new oral antidiabetic agent that enhances pancreatic islet cell responsiveness to glucose. An extensive clinical program involving approximately 22,000 patients and 7000 patient-years of exposure to vildagliptin has shown that the agent is well tolerated and efficacious in improving glycemic control in patients with type 2 diabetes mellitus (T2DM). Monotherapy trials have shown that significant HbA1c lowering is accompanied by body weight-neutral and lipid-neutral effects, low risk of edema, and low risk of hypoglycemia. These characteristics make vildagliptin a favorable partner for combination therapy. Studies of vildagliptin as an add-on to metformin have shown significant improvements in glycemic control (comparable to that of thiazolidinedione add-on), with the combination being well tolerated and associated with low risks for hypoglycemia and adverse effects on weight or lipid levels. Good tolerability and clinically relevant improvements in glycemic control have also been observed with vildagliptin as an add-on treatment to sulfonylurea, thiazolidinedione, or insulin treatment or in initial combination treatment with pioglitazone. Improved beta-cell function and glycemic control have been shown with vildagliptin in subjects with impaired glucose tolerance and in T2DM patients with mild hyperglycemia, with some evidence in the latter suggesting the potential for modifying disease course.

Combined thiazolidinedione–metformin treatment synergistically improves insulin signalling to insulin receptor substrate-1-dependent phosphatidylinositol 3-kinase, atypical protein kinase C and protein kinase B/Akt in human diabetic muscle

Insulin-stimulated glucose transport in muscle is impaired in type 2 diabetes, presumably reflecting reduced activation of atypical protein kinase C (aPKC) and protein kinase B (PKB/Akt). As previously shown, reductions in aPKC activation are seen at sub-maximal and maximal insulin stimulation, reductions in PKB activation are best seen at sub-maximal insulin stimulation and aPKC reductions at maximal insulin are partly improved by thiazolidinedione or metformin treatment. However, effects of combined thiazolidinedione-metformin treatment on aPKC or PKB activation by sub-maximal and maximal insulin are unknown. Type 2 diabetic patients were examined before and 5 to 6 weeks after combined thiazolidinedione-metformin therapy for activation of muscle aPKC and PKBbeta and their upstream activators, the insulin receptor (IR) and IRS-1-associated phosphatidylinositol 3-kinase (PI3K), during euglycaemic-hyperinsulinaemic clamp studies conducted with sub-maximal (400-500 pmol/l) and maximal (1400 pmol/l) insulin concentrations. Following combined thiazolidinedione-metformin therapy, increases in glucose disposal and increases in sub-maximal and maximal insulin-induced activities of all four muscle signalling factors, IR, IRS-1-dependent PI3K (IRS-1/PI3K), aPKC and PKBbeta, were observed. Increases in PKBbeta enzyme activity were accompanied by increases in phosphorylation of PKB and its substrate, AS160, which is needed for glucose transport. Despite improved aPKC activity, muscle aPKC levels, which are diminished in type 2 diabetes, were not altered. Combined thiazolidinedione-metformin treatment markedly improves sub-maximal and maximal insulin signalling to IR, IRS-1/PI3K, aPKC and PKBbeta in type 2 diabetic muscle. These improvements exceed those previously reported after treatment with either agent alone.

Myocardial heat shock protein 60 expression in insulin-resistant and diabetic rats

Heat shock protein 60 (HSPD1) plays a critical role in myocardial protection. Its reduced expression may lower myocardial protection against ischemic injury in the diabetic state. This study was conducted to investigate the natural course of fructose-fed insulin-resistant rats, define changes in myocardial HSPD1 expression, and determine the effects of thiazolidinedione or anti-hypertensive treatment. Results showed that insulin resistance with hyperinsulinemia and hypertension developed after 6 weeks of fructose feeding. This time-course study also showed that myocardial HSPD1 expression was mildly increased in week 6 (P=0.05) and significantly increased in week 8. Rosiglitazone-treated rats had restored systolic blood pressure (BP) and normalized plasma insulin level during oral glucose tolerance tests, whereas amlodipine-treated rats restored only systolic BP. Both amlodipine and rosiglitazone treatments normalized the abundance of myocardial HSPD1 expression in fructose-fed rats. When these rats received streptozotocin injection and diabetes developed, myocardial HSPD1 expression decreased despite persistent hypertension. In conclusion, this is the first study to report that myocardial HSPD1 expression is increased in high-fructose-fed rats, which may be due to increased BP. Once the high-fructose-fed rats developed diabetes with insulin deficiency, the myocardial HSPD1 expression decreased in spite of persistent hypertension.

Adiponectin secretion and response to pioglitazone is depot dependent in cultured human adipose tissue

The subcutaneous (S) and visceral (V) adipose tissue (AT) depots are increasingly recognized as distinct. To test the hypothesis that depot differences exist for adiponectin, fresh and cultured human VAT and SAT from obese type 2 diabetic (T2D) and obese nondiabetic (ND) subjects was examined to determine whether differences in adiponectin content and secretion occurred as a function of depot studied, diabetic status, and response to thiazolidinedione treatment. VAT and SAT were obtained by biopsy and AT explants cultured in defined media for 7 days. Protein expression was assessed by Western blot. Adiponectin content of conditioned medium was determined by radioimmunoassay. Diabetic status had no effect on adiponectin secretion over days 0-2 of culture. In ND SAT, secretion fell over days 2-4 but was sustained at greater levels vs. T2D SAT. In both ND and T2D VAT, adiponectin secretion was low, similar to T2D SAT. Over the 7-day culture period, cellular adiponectin increased in ND SAT and VAT; it remained unchanged in T2D SAT and VAT. Pioglitazone increased adiponectin secretion and content in all SAT. Pioglitazone failed to increase adiponectin secretion from either ND or T2D VAT and increased cellular content only in ND VAT. AT depot differences exist in the secretion of adiponectin and responsiveness to thiazolidinedione treatment. These data suggest that SAT, not VAT, appears to be the major contributor to increased circulating adiponectin levels in response to pioglitazone treatment.

Association between oral antidiabetic use, adverse events and outcomes in patients with type 2 diabetes

To quantify adverse events (AEs) associated with the use of metformin (MET), sulphonylureas (SUs) and thiazolidinediones (TZDs) in a usual care setting, and to assess the relationship of AEs to treatment patterns and glycaemic response in patients with type 2 diabetes. An electronic medical record database was used to identify patients with type 2 diabetes age >or=18 years from 1996 to 2005. Patients naïve to oral antidiabetic therapy were followed for 395 days postinitiation of MET, SU or TZD treatment. AEs related to study drugs were evaluated during the follow-up period. Baseline and follow-up A1C levels were compared by drug regimen. Associations between the change in A1C, drug regimen changes and AEs were evaluated. A total of 14,512 patients (mean age 60.8 years, 52.9% female) were identified. During the follow-up period, 12.7% of patients experienced an AE (8.6% MET, 15.9% SU and 19.8% TZD patients). SU and TZD patients were more likely to experience an AE than MET (p < 0.001) patients. AEs did not significantly influence A1C outcomes, although MET and SU patients experiencing an AE were more likely to add-on therapy (odds ratio (OR) = 1.34 and OR = 1.37, respectively; p < 0.05) than those without an AE. MET patients with AEs were more likely to switch therapy (OR = 1.91; p < 0.05) than those without an AE. The occurrence of AEs did not significantly impact glycaemic response to therapy. However, AEs may lead to greater treatment switches for patients receiving MET and add-on therapy for MET-treated and SU-treated patients.

Serum resistin is positively correlated with the accumulation of metabolic syndrome factors in type 2 diabetes

Resistin, secreted from adipocytes, causes insulin resistance in rodents. We reported that the G/G genotype of a resistin gene promoter single nucleotide polymorphism (SNP) at -420 increases type 2 diabetes (T2DM) susceptibility by enhancing promoter activity. We also showed that serum resistin was positively correlated with G at SNP-420, the duration of T2DM, and HbA1c in T2DM. The aim of this study was to determine the relation between serum resistin and factors related to the metabolic syndrome (MetS) in T2DM. We analysed 238 Japanese T2DM subjects (124 males and 114 females, age 60.2 +/- 11.3 years, body mass index (BMI) 24.1 +/- 3.9) whose overnight fasting sera were available. Serum resistin was measured using ELISA. Serum resistin was higher in subjects with either obesity (P = 0.041), low HDL (P = 0.004), high triglycerides (TG) (P = 0.019), hypertension (HT) (P = 0.001) or atherosclerosis (P = 0.012). Simple regression analysis revealed that serum resistin was correlated with lower HDL, TG and high-sensitivity C-reactive protein (hsCRP). Multiple regression analysis (or logistic regression analysis for HT), adjusted for age, gender, BMI and the duration of T2DM, revealed that serum resistin was correlated with lower HDL (P = 0.008), TG (P = 0.041), HT (P = 0.031) and hsCRP (P = 0.004). Serum resistin was positively correlated with the number of MetS factors, independent of age, gender and the duration of T2DM (P < 0.001). Adjustment by either thiazolidinedione (TZD) treatment or hsCRP had no effects on these findings. Serum resistin was positively correlated with the accumulation of MetS factors in T2DM.

Thiazolidinediones Downregulate Wnt/β-Catenin Signaling Via Multiple Mechanisms in Breast Cancer Cells

Thiazolidinediones (TZDs) have been demonstrated to possess antitumor effects in breast cancer cells, although the mechanisms are not well established. We sought to better define TZDs' antitumor effects and molecular mechanisms to permit rational utilization of these agents. We studied the effects of TZDs on DNA synthesis (BrdU enzyme-linked immunosorbent assay), gene expression (microarray, quantitative reverse transcription-polymerase chain reaction, and immunoblot), serine phosphorylation and localization of beta-catenin (nuclear/cytoplasmic fractionation and immunoblot), transactivation activity of beta-catenin/T cell factor 4 (TCF4) (luciferase assay with wild-type and mutant TCF4 responsive element), and beta-catenin/TCF4 complex (immunoprecipitation) in human breast cancer cells MDA-MB-231 and T47D. Troglitazone (TG) down-regulated DNA synthesis in MDA-MB-231 and T47D and decreased mRNA expression of the Wnt co-receptors frizzled-1 and low-density lipoprotein-related protein 6 (LRP6) as well as protein level of LRP6. TG also targets downstream Wnt signaling molecules in T47D cells. TG down-regulates p-beta-catenin (S33/S37/T41) and promotes translocation of beta-catenin into the nucleus. However, TG inhibits beta-catenin-mediated transactivation by down-regulating TCF4 protein levels, thereby inhibiting beta-catenin/TCF4 complex formation. Finally, we found that inhibition of Akt augments TG-mediated down-regulation of DNA synthesis and this result was accompanied with the decreased protein levels of Wnt signaling molecules: LRP6, beta-catenin, and TCF4. These results suggest that the Wnt/beta-catenin signaling pathway, which plays an important role in breast cancer pathogenesis, may be a target for TZD treatment. Moreover, a combination of TZDs and a specific Akt inhibitor may serve as a new approach to target Wnt/beta-catenin directly and via PI3K/Akt action on glycogen synthase-3beta.

The effects of rosiglitazone on aortic atherosclerosis of cholesterol-fed rabbits

Introduction Thiazolidinedione (TZD) is widely used a drug for the treatment of type 2 diabetes and protects against cardiovascular events in human. However, it is not clear whether TZD can directly inhibit the progression of atherosclerosis. To test the hypothesis whether administration of TZD could reduce the development of atherosclerosis, we studied the effects of rosiglitazone on aortic atherosclerosis of rabbits fed a cholesterol diet. Materials and methods Male Japanese White rabbits were fed a diet containing either 0.3% cholesterol diet (control group, n = 10) or 0.3% cholesterol with rosiglitazone (TZD-treated group, n = 12) for 16 weeks. We compared the plasma lipids and the extent of aortic atherosclerosis between two groups. Results and conclusions TZD treatment significantly resulted in the reduction of aortic atherosclerosis by 21% in the aortic arch (p < 0.01), 20% in the thoracic aorta (p = 0.14), and 28% in the abdominal aorta (p = 0.25), without affecting the plasma levels of triglycerides, total cholesterol, high-density lipoprotein cholesterol, glucose and insulin. Immunohistochemical staining showed that the cellular components (macrophages and smooth muscle cells) of the lesions of TZD-treated rabbits were unchanged compared to those of control rabbits. In addition, TZD treatment also led to dramatic improvement of fatty liver in cholesterol-fed rabbits. Our results suggest that the activation of PPARγ can be beneficial for the treatment of atherosclerosis and fatty liver independent upon the improvement of plasma lipids and glucose metabolism.