Pio Treatment Research Articles

Effect of parthenolide, an NLRP3 inflammasome inhibitor, on insulin resistance in high-fat diet-obese mice.

The activation of Nod-like receptor proteins (NLRP3) containing the pyrin domain inflammasome is a hallmark of the pathogenesis of metabolic disorders. Inhibition of the NLRP3 inflammasome by phytoconstituents has been attempted as a strategy to mitigate these disorders. Therefore, the present study aimed to evaluate the efficacy of an NLRP3 inflammasome inhibitor, parthenolide (PN; 5mg/kg i.p.) against inflammation and insulin resistance in high-fat diet (HFD) - obese mice. Treatment with PN and pioglitazone (PIO; 30mg/kg p.o.) attenuated lipopolysaccharide (LPS; 1 ng/ml) - induced elevation of tumor necrosis factor-α and interleukin-1β in mouse peritoneal macrophages in a dose-dependent manner. Sixty days of PN and PIO treatment marginally reduced obesity-induced insulin resistance in HFD-obese mice. PN treatment also decreased blood glucose from 14th to 60th day, supporting the hypothesis of simultaneous attenuation of inflammation and insulin resistance in obese mice. Thus, PN treatment was also evident with significant improvement in glucose tolerance and peripheral insulin resistance validated through the respective tolerance tests. Therefore, the present study suggests that PN, an NLRP3 inflammasome inhibitor, could be a possible therapeutic agent for attenuating obesity-induced insulin resistance.

Role of peroxisome proliferators-activated receptor-gamma in advanced glycation end product-mediated functional loss of voltage-gated potassium channel in rat coronary arteries

BackgroundHigh blood glucose impairs voltage-gated K+ (Kv) channel-mediated vasodilation in rat coronary artery smooth muscle cells (CSMCs) via oxidative stress. Advanced glycation end product (AGE) and receptor for AGE (RAGE) axis has been found to impair coronary dilation by reducing Kv channel activity in diabetic rat small coronary arteries (RSCAs). However, its underlying mechanism remain unclear. Here, we used isolated arteries and primary CSMCs to investigate the effect of AGE incubation on Kv channel-mediated coronary dilation and the possible involvement of peroxisome proliferators-activated receptor (PPAR) -γ pathway.MethodsThe RSCAs and primary CSMCs were isolated, cultured, and treated with bovine serum albumin (BSA), AGE-BSA, alagrebrium (ALA, AGE cross-linking breaker), pioglitazone (PIO, PPAR-γ activator) and/or GW9662 (PPAR-γ inhibitor). The groups were accordingly divided as control, BSA, AGE, AGE + ALA, AGE + PIO, or AGE + PIO + GW9662. Kv channel-mediated dilation was analyzed using wire myograph. Histology and immunohistochemistry of RSCAs were performed. Western blot was used to detect the protein expression of RAGE, major Kv channel subunits expressed in CSMCs (Kv1.2 and Kv1.5), PPAR-γ, and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase-2 (NOX-2).ResultsAGE markedly reduced Forskolin-induced Kv channel-mediated dilation of RSCAs by engaging with RAGE, and ALA or PIO significantly reversed the functional loss of Kv channel. In both RSCAs and CSMCs, AGE reduced Kv1.2/1.5 expression, increased RAGE and NOX-2 expression, and inhibited PPAR-γ expression, while ALA or PIO treatment partially reversed the inhibiting effects of AGE on Kv1.2/1.5 expression, accompanied by the downregulation of RAGE and decreased oxidative stress. Meanwhile, silencing of RAGE with siRNA remarkably alleviated the AGE-induced downregulation of Kv1.2/1.5 expression in CSMCs.ConclusionAGE reduces the Kv channel expression in CSMCs and further impairs the Kv channel-mediated dilation in RSCAs. The AGE/RAGE axis may enhance oxidative stress by inhibiting the downstream PPAR-γ pathway, thus playing a critical role in the dysfunction of Kv channels.

Pioglitazone improves porcine oocyte maturation and subsequent parthenogenetic embryo development in vitro by increasing lipid metabolism.

Optimization of culture conditions is important to improve oocyte maturation and subsequent embryo development. In particular, this study analyzed the effects of increasing concentrations of PIO in the maturation medium on spindle formation and chromosome alignment, glutathione, and intracellular ROS levels and expression of selected genes related to maternal markers, apoptosis, and lipid metabolism. The percentage of oocytes displaying normal spindle formation and chromosome alignment was higher in the 1 µM PIO (1 PIO)-treated group than in the control group. The glutathione level was significantly higher in the 1 PIO-treated group than in the control group, while the reactive oxygen species level did not differ. Expression of maternal marker (MOS and GDF9), antiapoptotic (BIRC5), and lipid metabolism-related (ACADS, CPT2, SREBF1, and PPARG) genes was higher in the 1 PIO-treated group than in the control group, while expression of a proapoptotic gene (CASP3) was lower. The blastocyst formation rate and the percentage of blastocysts that reached at least the hatching stage on Days 6 and 7, and the percentage of blastocysts containing more than 128 cells were significantly higher in the 1 PIO-treated group than in the control group. These results indicate that PIO treatment during in vitro maturation improves porcine oocyte maturation and subsequent parthenogenetic embryo development mainly by enhancing lipid metabolism and antioxidant defense in oocytes.

Activated PPARγ Abrogates Misprocessing of Amyloid Precursor Protein, Tau Missorting and Synaptotoxicity.

Type 2 diabetes increases the risk for dementia, including Alzheimer’s disease (AD). Pioglitazone (Pio), a pharmacological agonist of the peroxisome proliferator-activated receptor γ (PPARγ), improves insulin sensitivity and has been suggested to have potential in the management of AD symptoms, albeit through mostly unknown mechanisms. We here investigated the potential of Pio to counter synaptic malfunction and loss, a characteristic of AD pathology and its accompanying cognitive deficits. Results from experiments on primary mouse neuronal cultures and a human neural cell line (SH-SY5Y) show that Pio treatment attenuates amyloid β (Aβ)-triggered the pathological (mis-) processing of amyloid precursor protein (APP) and inhibits Aβ-induced accumulation and hyperphosphorylation of Tau. These events are accompanied by increased glutamatergic receptor 2B subunit (GluN2B) levels that are causally linked with neuronal death. Further, Pio treatment blocks Aβ-triggered missorting of hyperphosphorylated Tau to synapses and the subsequent loss of PSD95-positive synapses. These latter effects of Pio are PPARγ-mediated since they are blocked in the presence of GW9662, a selective PPARγ inhibitor. Collectively, these data show that activated PPARγ buffer neurons against APP misprocessing, Tau hyperphosphorylation and its missorting to synapses and subsequently, synaptic loss. These first insights into the mechanisms through which PPARγ influences synaptic loss make a case for further exploration of the potential usefulness of PPARγ agonists in the prevention and treatment of synaptic pathology in AD.

Pioglitazone's beneficial effects on erectile function preservation after cavernosal nerve injury in the rat are negated by inhibition of the insulin-like growth factor-1 receptor: a preclinical study.

To determine if the insulin-like growth factor-1 (IGF-1) pathway is involved in the improvement in erectile function recovery in rats after nerve crush injury treated with pioglitazone (Pio). Sprague-Dawley rats were divided into four groups. The first group received sham operation (n = 5). The second group underwent bilateral cavernous nerve injury (BCNI, n = 7). The third group received BCNI and Pio treatment (BCNI + Pio, n = 7), whereas the fourth group underwent BCNI with Pio treatment and IGF-1 inhibition (BCNI + Pio + JB-1, n = 7). The IGF-1 receptor (IGF-1R) was inhibited by JB-1, a small molecular antagonist of the receptor. After 14 days of treatment, erectile function was measured via intracorporal pressure normalized to mean arterial pressure (ICP/MAP) and the major pelvic ganglion and cavernous nerve harvested for western blot and immunohistochemistry (IHC) of phosphorylated-IGF-1Rβ (p-IGF-1Rβ), phosphorylated-ERK1/2 (p-ERK1/2), and neuronal NOS (nNOS). BCNI + Pio animals exhibited improvements in ICP/MAP, similar to Sham animals, and BCNI + Pio + JB-1 rats demonstrated a reduced ICP/MAP similar to BCNI-only rats at all measured voltages. Western blot results showed upregulation of p-IGF-1Rβ was observed in the BCNI + Pio group. Low levels of p-ERK1/2 were seen in the JB-1-treated animals. The immunoblot results were supported by IHC findings. Intense IHC staining of nNOS was detected in the BCNI + Pio group. The group treated with JB-1 showed minimal protein expression of p-ERK1/2, nNOS, and p-IGF-1Rβ. Pio improves erectile function in rats undergoing BCNI via an IGF-1-mediated pathway.

Pioglitazone ameliorates retinal ischemia/reperfusion injury via suppressing NLRP3 inflammasome activities.

To explore the role of Pioglitazone (Pio) on a mouse model of retinal ischemia/reperfusion (I/R) injury and to elucidate the potential mechanism. Retinal ischemia was induced in mice by increasing the intraocular pressure, and Pio was administered 4h though periocular injection before I/R. The number of cells in the ganglion cell layer (GCL) was counted 7d after retinal I/R injury. Glial fibrillary acidic protein (GFAP), nuclear factor-kappa B (NF-κB), p38, phosphorylated-p38, PPAR-γ, interleukin-1β (IL-1β), Toll-like receptor 4 (TLR4), NLRP3, cleaved caspase-1, caspase-1 were determined by real-time polymerase chain reaction and Western blotting. Pio promoted the survival of retinal cells in GCL following retinal I/R injury (P<0.05). Besides, retinal I/R injury stimulated the expression of GFAP and TLR4, which were partially reversed by Pio treatment (P<0.05). Retinal I/R injury-upregulated expression of NLRP3, cleaved caspase-1, IL-1β was attenuated after Pio treatment (P<0.05). Moreover, I/R injury induced activation of NF-κB and p38 were inhibited by Pio treatment (P<0.05). Pio promotes retinal ganglion cells survival by suppressing I/R-induced activation of TLR4/NLRP3 inflammasomes via inhibiting NF-κB and p38 phosphorylation.

Pioglitazone stabilizes atherosclerotic plaque by regulating the Th17/Treg balance in AMPK-dependent mechanisms

BackgroundPioglitazone (PIO), a thiazolidinediones drug, is a well-known anti-diabetic medicine, but its anti-atherosclerotic effects remain controversial. Thus it is important to investigate the effects of PIO on atherogenesis and the relevant mechanisms.MethodsFor in vitro studies, primary cultured or AMP-activated protein kinase (AMPK) inhibited splenocytes were treated with oxidized low density lipoprotein (ox-LDL) or ox-LDL plus PIO. Percentage of T helper 17 (Th17) and regulatory T (Treg) cells were determined by flow cytometry. Expression of AMPK, interleukin-17 (IL-17) and forkhead box P3 (FoxP3) were detected by Western blots. For in vivo studies, apolipoprotein E–deficient (apoE−/−) mice fed with western diet were treated with PIO or vehicle for 8 weeks respectively. Percentage of Th17 and Treg cells in spleen were measured by immunohistochemical analysis. The atherosclerotic lesions were analyzed using oil red O staining, and collagen types I and III in atherosclerotic lesions were stained by Sirius red. Expression of IL-17 and FoxP3 were determined by quantitative polymerase chain reaction.ResultsIn cultured primary splenocytes, PIO dramatically inhibited Th17 and raised Treg. Intriguingly, pharmacological and genetic AMPK inhibitions abolished PIO-induced Treg elevation and Th17 inhibition. Moreover, PIO significantly induced AMPK phosphorylation, decreased IL-17+ and increased FoxP3+ cells in spleen of apoE−/− mice. Finally, PIO did not alter plaque area, but intriguingly, stabilized atherosclerotic plaque through collagen induction in apoE−/− mice. PIO treatment also improved Th17/Treg balance in atherosclerotic lesions.ConclusionsPIO exhibits anti-atherosclerotic effects for stabilization of atherosclerotic plaque through regulating the Th17/Treg balance in an AMPK-dependent manner.

P 189 - PPAR gamma activation can improve aberrant redox regulation in hypertension

Dysregulation of redox- sensitive signaling plays an important role in develompment of hypertension. PPAR gamma belongs to nuclear receptors and is a potent nutritional sensor and redox regulator. In our study we focused on the role of PPAR gamma activation of RAS/ROS regulation, and antioxidant response in spontaneously hypertensive rats (SHR). Treatment was performed by gavage with PPAR gamma agonist pioglitazone (PIO - 10 mg/kg/day, 10 days). Treatment with PPAR gamma agonist PIO in influenced blood pressure, redox- sensitive responses, lipid profile and homocystein level in age- dependent manner. The improvement parameters in hypertension and redox regulation have been observed in young animals, but not in adult SHR rats. PIO treatment influenced redox regulation (gene expression of Nrf2 and SOD isoforms) correlated with SOD a CAT activity and NO bioavailibility. PPAR gamma activation has been corelated with downregulation of and AT1R - RAS axis and „up-regulation“ of AT2R and Mas receptor. This most sensitive tissue responses in RAS/ROS signaling and NO level have been found primary in kidney of young hypertensive rats.

Pioglitazone-induced improvements in insulin sensitivity occur without concomitant changes in muscle mitochondrial function

AimsPioglitazone (Pio) is known to improve insulin sensitivity in skeletal muscle. However, the role of Pio in skeletal muscle lipid metabolism and skeletal muscle oxidative capacity is not clear. The aim of this study was to determine the effects of chronic Pio treatment on skeletal muscle mitochondrial activity in individuals with type 2 diabetes (T2D). Materials and MethodsTwenty-four participants with T2D (13M/11F 53.38±2.1years; BMI 36.47±1.1kg/m2) were randomized to either a placebo (CON, n=8) or a pioglitazone (PIO, n=16) group. Following 12weeks of treatment, we measured insulin sensitivity by hyperinsulinemic–euglycemic clamp (clamp), metabolic flexibility by calculating the change in respiratory quotient (ΔRQ) during the steady state of the clamp, intra- and extra-myocellular lipid content (IMCL and EMCL, respectively) by 1H magnetic resonance spectroscopy (1H-MRS) and muscle maximal ATP synthetic capacity (ATPmax) by 31P-MRS. ResultsFollowing 12weeks of PIO treatment, insulin sensitivity (p<0.0005 vs. baseline) and metabolic flexibility (p<0.05 vs. CON) significantly increased. PIO treatment significantly decreased IMCL content and increased EMCL content in gastrocnemius, soleus and tibialis anterior muscles. ATPmax was unaffected by PIO treatment. ConclusionsThese results suggest that 12weeks of pioglitazone treatment improves insulin sensitivity, metabolic flexibility and myocellular lipid distribution without any effect on maximal ATP synthetic capacity in skeletal muscle. Consequently, pioglitazone-induced enhancements in insulin responsiveness and fuel utilization are independent of mitochondrial function.

Abstract 657: PDGF versus Fructose Dysregulation of Insulin Receptor Signaling in Vascular Smooth Muscle Cells: Pioglitazone-mediated Sensitization of Insulin Receptor Signaling is Associated with Diminished Vascular Smooth Muscle Cell Proliferation

Previous studies have shown that PDGF-induced activation of mTOR/p70S6kinase signaling is associated with dysregulation of insulin receptor substrates (IRS1 and IRS2) in vascular smooth muscle cells (VSMCs). In addition, pioglitazone (PIO, an antidiabetic drug) has been shown to inhibit mTOR/p70S6kinase signaling thereby suppressing PDGF-induced VSMC proliferation. The role of IRS1/IRS2 on proliferative signaling events has not yet been examined in VSMCs treated with fructose (an agent that induces insulin resistance) or PIO (an insulin sensitizer). In the present study, unlike PDGF treatment (48 hr) that enhanced IRS serine phosphorylation and diminished IRS2 expression, exposure of human aortic VSMCs to D-fructose (5.5 to 25 mM) for 48 hr resulted in a concentration-dependent increase in IRS1 serine phosphorylation (up to ~5-fold , n = 3), but without any changes in IRS2 protein expression. In D-fructose-treated VSMCs, Akt phosphorylation in response to acute insulin exposure (6 min) was markedly decreased (~69%, n = 3), suggesting that IRS1 serine phosphorylation alone is sufficient to dysregulate insulin receptor signaling. In addition, D-fructose treatment led to a further increase in PDGF-induced VSMC proliferation (~40%, n = 3). Furthermore, we examined the effects of PIO on PDGF-induced dysregulation of IRS in VSMCs. Notably, PIO treatment (30 μM, 48 hr) by itself led to an upregulation of IRS-2 expression (~2.5-fold) and also diminished PDGF-induced IRS1 serine phosphorylation (~63%). Under these conditions, PIO treatment resulted in significant inhibition of PDGF-induced cyclin D1 expression, Rb phosphorylation, and VSMC proliferation (~83%, n = 3). In conclusion, the present study reveals that dysregulated insulin signaling (by PDGF and fructose) is associated with enhanced VSMC proliferation, whereas insulin sensitization (by PIO) is associated with inhibition of VSMC proliferation. Together, these findings underscore the significance of intact insulin signaling toward limiting exaggerated VSMC proliferation in the vessel wall.

Pioglitazone alters monocyte populations and stimulates recent thymic emigrants in the BBDZR/Wor type 2 diabetes rat model

BackgroundType 2 diabetes is commonly characterized by insulin deficiency and decreased sensitivity of insulin receptors, leading to a chronic state of hyperglycemia in individuals. Disease progression induces changes in the immune profile that engenders a chronic inflammatory condition. Thiazolidinedione (TDZ) drugs, such as Pioglitazone (Pio), aid in controlling disease symptoms. While the mechanisms by which Pio controls hyperglycemia are beginning to be understood, relatively little is known about the effects of Pio on suppression of the systemic immune phenotype, attributed to visceral adipose tissue and macrophages.MethodsHere, we utilize the recently developed BBDZR/Wor type 2 diabetes rat model to test our hypothesis that a selective in vivo growth of CD3+T cells in the spleen contributes to the increase in T lymphocytes, including Tregs, independent of visceral adipose tissue. We investigated the systemic effects of Pio on multifactorial aspects of the disease-induced immune phenotype both in vivo and in vitro in normal, non-diabetic animals and in disease.ResultsOur work revealed that Pio reversed the lymphopenic status of diabetic rats, in part by an increase in CD3+ T lymphocytes and related subsets. Moreover, we found evidence that Pio caused a selective growth of newly differentiated T lymphocytes, based on the presence of recent thymic emigrants in vivo. To investigate effects of Pio on the inflammatory milieu, we examined the production of the signature cytokines TNF-α and IL-1β and found they were reduced by Pio-treatment, while the levels of IL-4, an anti-inflammatory mediator, were significantly increased in a Pio-dependent manner. The increase in IL-4 production, although historically attributed to macrophages from visceral adipose tissue under other conditions, came also from CD3+ T lymphocytes from the spleen, suggesting splenocytes contribute to the Pio-induced shift towards an anti-inflammatory phenotype.ConclusionsWe show for the first time that Pio treatment significantly suppresses the systemic inflammatory status in the BBDZR/Wor type 2 diabetes rat model by the selective growth of newly differentiated CD3+ T cells and by increasing CD3+IL-4 production in immigrant spleen lymphocytes.

Pioglitazone inhibits the secretion of proinflammatory cytokines and chemokines in astrocytes stimulated with lipopolysaccharide.

Neuroinflammation caused by the secretion of cytokines and chemokines by glial cells can promote the development of neurodegenerative disorders. The aim of this study was to explore the effects of pioglitazone (Pio), a drug that induces release of inflammatory mediators, on cytokine and chemokine secretion in astrocytes stimulated with lipopolysaccharide (LPS) to induce inflammation. Astrocytes obtained from the cerebral cortex of newborn C57BL/6 mice and grown in culture were stimulated with LPS and treated with Pio. Treatment of astrocytes with LPS significantly increased the levels of pro-inflammatory factors nitric oxide (NO), tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-6, and IL-8, but decreased the level of anti-inflammatory factors IL-4 and IL-10 (p < 0.05) compared to untreated control astrocytes. Pio treatment in LPS-stimulated astrocytes had the opposite effect, inhibiting secretion of NO, TNF-α, IL-1β, IL-6, and IL-8 and enhancing IL-4 and IL-10 secretion (p < 0.05). In addition, Pio treatment suppressed the expression of pro-inflammatory chemokines Ccl20, Mcp-1, and Mip-1α mRNA in astrocytes stimulated with LPS (p < 0.05). These results showed that Pio can inhibit the neuroinflammatory response in LPS-activated astrocytes and this response may result from the regulation of cytokine and chemokine secretion from astrocytes.

Inhibitory effects of peroxisome proliferator-activated receptor γ agonists on collagen IV production in podocytes.

Peroxisome proliferator-activated receptor-γ (PPAR-γ) agonists have beneficial effects on the kidney diseases through preventing microalbuminuria and glomerulosclerosis. However, the mechanisms underlying these effects remain to be fully understood. In this study, we investigate the effects of PPAR-γ agonist, rosiglitazone (Rosi) and pioglitazone (Pio), on collagen IV production in mouse podocytes. The endogenous expression of PPAR-γ was found in the primary podocytes and can be upregulated by Rosi and Pio, respectively, detected by RT-PCR and Western blot. PPAR-γ agonist markedly blunted the increasing of collagen IV expression and extraction in podocytes induced by TGF-β. In contrast, adding PPAR-γ antagonist, GW9662, to podocytes largely prevented the inhibition of collagen IV expression from Pio treatment. Our data also showed that phosphorylation of Smad2/3 enhanced by TGF-β in a time-dependent manner was significantly attenuated by adding Pio. The promoter region of collagen IV gene contains one putative consensus sequence of Smad-binding element (SBE) by promoter analysis, Rosi and Pio significantly ameliorated TGF-β-induced SBE4-luciferase activity. In conclusion, PPAR-γ activation by its agonist, Rosi or Pio, in vitro directly inhibits collagen IV expression and synthesis in primary mouse podocytes. The suppression of collagen IV production was related to the inhibition of TGF-β-driven phosphorylation of Smad2/3 and decreased response activity of SBEs of collagen IV in PPAR-γ agonist-treated mouse podocytes. This represents a novel mechanistic support regarding PPAR-γ agonists as podocyte protective agents.

Pioglitazone treatment enhances the sympathetic nervous system response to oral carbohydrate load in obese individuals with metabolic syndrome

ContextInsulin resistance is associated with blunted sympathetic nervous system (SNS) response to carbohydrate ingestion which may contribute to postprandial hypotension and impaired body weight homeostasis. ObjectiveThis study was conducted to examine the effects of pharmacological insulin sensitization on whole-body norepinephrine kinetics during a standard 75-g oral glucose tolerance test (OGTT) in obese, insulin resistant subjects with metabolic syndrome. MethodsUn-medicated individuals (n=42, mean age 56±0.8yrs, body mass index 34±0.6kg/m2) were randomised to 12-weeks pioglitazone (PIO, 15mg for 6weeks, then 30mg daily) or placebo using a double-blind, parallel group design. Whole-body norepinephrine kinetics (arterial norepinephrine concentration, calculated spillover and clearance rates), spontaneous cardiac baroreflex sensitivity, heart rate and blood pressure were measured at times 0, 30, 60, 90 and 120minutes during OGTT. Insulin sensitivity was assessed by euglycemic hyperinsulinemic clamp (M) and Matsuda index. ResultsPIO increased clamp derived glucose utilisation by 35% (P<0.001) and there were concurrent reductions in inflammatory status and plasma triglycerides (P<0.05). Fasting norepinephrine kinetic parameters were unaltered. PIO treatment was associated with lower plasma insulin incursions, greater reduction in diastolic blood pressure and enhanced baroreflex sensitivity during OGTT (P all <0.05). The overall norepinephrine spillover response (AUC0–120) increased significantly in the PIO group (group × time interaction, P=0.04), with greatest increment at 30minutes post-glucose (101±38ng/min at baseline versus 241±48ng/min post treatment, P=0.04) and correlated with percent improvement in M. ConclusionsPIO enhances the early postprandial SNS response to carbohydrate ingestion.

EFFECT OF PIOGLITAZONE ON HEPATIC ULTRA-STRUCTURE AND METABOLIC CHANGES INDUCED BY A HIGH SUCROSE DIET IN RATS

Pioglitazone (Pio) is a PPAR-γ agonist insulin sensitizer has anti-inflammatory activity. Our novel aspect was to investigate its hepatic anti-inflammatory activity at the level of ultra-structure and enzymatic changes in a high sucrose diet animal model. Forty male Sprague Dawley rats were divided into four equal groups: Control; control Pio; high sucrose diet; high sucrose diet Pio treated groups. Fourteen weeks later, serum glucose, insulin, lipogram, gamma glutamyle transferase, alanine transaminase, aspartate transaminase, alkaline phosphatase, fetuin-A and adiponectin levels were measured. Hepatic tissue homogenate levels of tumor necrosis factor-α, interleukin-6 and myeloperoxidase activity were determined. Microscopic and ultra-structure hepatic changes were conducted in all animal groups. Administration of Pio in HS+Pio group reduced significantly the hepatic inflammatory markers and the hepatic enzymes compared with HS group. Both light and electron microscopic examination revealed a great improvement of the hepatic tissue with Pio treatment. This study suggested that Pio treatment in obesity; in addition to insulin sensitizing activity; could protect the liver from the development of hepatic inflammation induced by a high sucrose diet not only at the enzymatic but also at ultra-structure levels.

Preventive roles of swimming exercise and pioglitazone treatment on hepatic dysfunction in a rat model of metabolic syndrome

Pioglitazone (Pio) and swimming exercise (SE) are insulin sensitisers. This investigation was suggested because of the significant side effects associated with Pio treatment in metabolic syndrome (MetS). This study was, therefore, designed to investigate the preventive role of Pio treatment and SE in terms of efficiency and pathological changes in MetS in a rat model. Sixty male Sprague-Dawley rats were distributed equally among 6 groups: (i) control group (C), (ii) exercised control group (C+E), (iii) Pio-treated control group (C+Pio), (iv) group with MetS, (v) group with MetS treated with Pio (MetS+Pio), and (vi) exercised MetS group (MetS+E). Systolic blood pressure and heart rate were measured at the end of the experiments (16 weeks). Retro-orbital blood samples were used to determine the serum levels of glucose, insulin, lipids, gamma glutamyl transferase, alanine transaminase, aspartate transaminase, alkaline phosphatase, fetuin-A, and adiponectin. Semiquantitative reverse transcriptase - PCR insulin gene expression assays and hepatic histopathological examination were conducted. Swimming exercise significantly improved all of the aforementioned parameters, more so than the Pio treatment. In particular, the serum hepatic enzyme levels and hepatic histopathological changes were improved compared with the MetS group. These results suggested that swimming exercise might be an alternative physiological preventive tool against hepatic dysfunction to avoid the side effects associated with Pio treatment, and this could be demonstrated in a rat model of metabolic syndrome.

Divergent compensatory responses to high-fat diet between C57BL6/J and C57BLKS/J inbred mouse strains

Impaired glucose tolerance (IGT) and type 2 diabetes (T2DM) are polygenic disorders with complex pathophysiologies; recapitulating them with mouse models is challenging. Despite 70% genetic homology, C57BL/6J (BL6) and C57BLKS/J (BLKS) inbred mouse strains differ in response to diet- and genetic-induced obesity. We hypothesized these differences would yield insight into IGT and T2DM susceptibility and response to pharmacological therapies. To this end, male 8-wk-old BL6 and BLKS mice were fed normal chow (18% kcal from fat), high-fat diet (HFD; 42% kcal from fat), or HFD supplemented with the PPARγ agonist pioglitazone (PIO; 140 mg PIO/kg diet) for 16 wk. Assessments of body composition, glucose homeostasis, insulin production, and energy metabolism, as well as histological analyses of pancreata were undertaken. BL6 mice gained weight and adiposity in response to HFD, leading to peripheral insulin resistance that was met with increased β-cell proliferation and insulin production. By contrast, BLKS mice responded to HFD by restricting food intake and increasing activity. These behavioral responses limited weight gain and protected against HFD-induced glucose intolerance, which in this strain was primarily due to β-cell dysfunction. PIO treatment did not affect HFD-induced weight gain in BL6 mice, and decreased visceral fat mass, whereas in BLKS mice PIO increased total fat mass without improving visceral fat mass. Differences in these responses to HFD and effects of PIO reflect divergent human responses to a Western lifestyle and underscore the careful consideration needed when choosing mouse models of diet-induced obesity and diabetes treatment.

Inflammatory cytokines and chemokines, skeletal muscle and polycystic ovary syndrome: Effects of pioglitazone and metformin treatment

ObjectiveChronic low-grade inflammation is a common feature of insulin resistant states, including obesity and type 2 diabetes. Less is known about inflammation in Polycystic Ovary Syndrome (PCOS). Thus we evaluated the impact of PCOS on circulating cytokine levels and the effects of anti-diabetic therapies on insulin action, cytokine and chemokine levels and inflammatory signaling in skeletal muscle. MethodsTwenty subjects with PCOS and 12 healthy normal cycling (NC) subjects of similar body mass index were studied. PCOS subjects received oral placebo or pioglitazone, 45mg/d, for 6months. All PCOS subjects then had metformin, 2g/day, added to their treatment. Circulating levels of cytokines, chemokines, and adiponectin, skeletal muscle markers of inflammation and phosphorylation of signaling proteins, insulin action evaluated by the hyperinsulinemic/euglycemic clamp procedure and Homeostasis Model Assessment of Insulin Resistance were measured. ResultsCirculating levels of a number of cytokines and chemokines were generally similar between PCOS and NC subjects. Levels in PCOS subjects were not altered by pioglitazone or metformin treatment, even though whole body insulin action and adiponectin levels increased with pioglitazone. In spite of the lack of change in levels of cytokines and chemokines, several markers of inflammation in skeletal muscle were improved with Pio treatment. ConclusionsPCOS may represent a state of elevated sensitivity of inflammatory cells in skeletal muscle to cytokines and chemokines, a property that could be reversed by pioglitazone treatment together with improved insulin action.

The Role of Peroxisome Proliferator-Activated Receptor and Effects of Its Agonist, Pioglitazone, on a Rat Model of Optic Nerve Crush: PPARγ in Retinal Neuroprotection

It has been shown that peroxisome proliferators-activated receptor gamma (PPARγ) is beneficial for central nervous system injury. However its role on optic nerve injury remains unknown. In the present study, we examined the change of PPARγ expression in rat retina following optic nerve injury and investigated the effect of pioglitazone (Pio), a PPARγ agonist, on retinal ganglion cells (RGCs) neuroprotection using a rat optic nerve crush (ONC) model. Our results showed that PPARγ mRNA and protein levels were increased after ONC, and most of PPARγ-immunoreactive cells colocalized with Müller cells. Pio treatment significantly enhanced the number of surviving RGCs and inhibited RGCs apoptosis induced by ONC. However, when PPARγ antagonist GW9662 was used, these neuroprotective effects were abolished. In addition, pio attenuated Müller cell activation after ONC. These results indicate that PPARγ appears to protect RGCs from ONC possibly via the reduction of Müller glial activation. It provides evidence that activation of PPARγ may be a potential alternative treatment for RGCs neuroprotection.

P4‐258: Genome‐wide screen for variants that modify the known relationship between cerebrospinal fluid Beta‐Amyloid 42 and ptau levels

suggests that PIO improves or protects neuronal mitochondrial function and increases mitochondrial biogenesis in vivo and in vitro. Additionally, results of behavioral studies indicate that TZD treatment may ameliorate some of the cognitive deficits in murine models of Alzheimer’s disease (AD) and human AD. This initial exploration examined the effects of low-dose PIO treatment on brain functional connectivity in awake rats using fMRI. Methods: Awake, adult male Wistar rats were acclimated to the scanning procedure for approximately 7 days. A baseline scan was performed 2.53h after dosing with vehicle (citric acid) using a 4.7 Tesla/30cm horizontal magnet and blood-oxygen-level-dependent (BOLD) imaging. Thereafter, rats were dosed orally with vehicle or 1 of 4 dosages of PIO (0.04, 0.08, 0.16, 0.32 mg/kg). One group of vehicleand one group of PIO-treated rats were scanned after the 2nd dosing; all rats were scanned after the last (7th) daily dose (n1⁄45 for each group). Results: For region of interest (ROI) analysis, each brain was demarcated into 57 ROIs and cross-correlation coefficients between pairs of ROIs were calculated to evaluate functional connectivity between ROIs. Seventeen ROI pairs showed significant (P<0.005, uncorrected) changes in functional connectivity on Day 7 of treatment with PIO compared with vehicle. Significant changes were seen in two pairs of ROIs on day 2 of dosing with PIO compared to the control group. Using the CA1 region of the hippocampus as a seed region, a voxel-by-voxel analysis indicated increased connectivity between the seed and the hypothalamus and ventral thalamus after 7 days of dosing. Conclusions: Our results indicate that administration of pioglitazone at doses lower than those used to treat T2D leads to changes in functional connectivity in conscious rats, as assessed by fMRI BOLD imaging; some changesmay occur within 2 days of treatment. These results provide preliminary evidence that low doses of PIO lead to changes in the brain and support the hypothesis that this drug may be a beneficial pharmacotherapy for neurodegenerative diseases such as AD.