Pentoxifylline has anti-inflammatory properties and could suppress some inflammatory processes including tumor necrosis factor-alpha (TNF-α) production. We assessed the effects of a two-month administration of pentoxifylline on nuclear factor-kappa B (NFκB) pathways in patients with coronary artery disease (CAD) in which inflammatory pathways, especially NFκB transcription factors, have a critical role.
A double-blind randomized placebo-controlled study design was used. Forty CAD patients were randomized to either 2 months of pentoxifylline treatment (1200 mg/day) (n = 20) or placebo treatment (n = 20). Blood samples were obtained just before and after two months of treatment. P50 protein concentration in peripheral blood mononuclear cells (PBMCs) was measured by Enzyme Linked ImmunoSorbent Assay (ELISA) method.
P50 concentration did not significantly change during two months of pentoxifylline administration.
Longer pentoxifylline administration is needed to see its favorable effects on NFκB family elements.
Coronary Artery Diseases; Inflammation; NFκB; Pentoxifylline
There are many sources of nutritionally mediated oxidative stress that trigger inflammatory cascades along short and long time frames. These events are primarily mediated via NFκB. On the short-term scale postprandial inflammation is characterized by an increase in circulating levels of IL-6 and TNF-α and is mirrored on the long-term by proinflammatory gene expression changes in the adipocytes and peripheral blood mononuclear cells (PBMCs) of obese individuals. Specifically the upregulation of CCL2/MCP-1, CCL3/MIP-1α, CCL4/MIP-1β, CXCL2/MIP-2α, and CXCL3/MIP-2β is noted because these changes have been observed in both adipocytes and PBMC of obese humans. In comparing numerous human intervention studies it is clear that pro-inflammatory and anti-inflammatory consumption choices mediate gene expression in humans adipocytes and peripheral blood mononuclear cells. Arachidonic acid and saturated fatty acids (SFAs) both demonstrate an ability to increase pro-inflammatory IL-8 along with numerous other inflammatory factors including IL-6, TNFα, IL-1β, and CXCL1 for arachidonic acid and IGB2 and CTSS for SFA. Antioxidant rich foods including olive oil, fruits, and vegetables all demonstrate an ability to lower levels of IL-6 in PBMCs. Thus, dietary choices play a complex role in the mediation of unavoidable oxidative stress and can serve to exacerbate or dampen the level of inflammation.
Background. This study was performed to investigate the influence of a short-term treatment with pioglitazone versus placebo on inflammatory activation of mononuclear cells (mRNA expression/protein secretion of inflammatory markers).
Methods and Results. Sixty-three patients with well-controlled type 2 diabetes (52 males, 11 females, age (Mean ± SD): 66 ± 7 yrs, disease duration: 6.6 ± 9.6 yrs, HbA1c: 6.7 ± 0.6%) were randomized to additional 45 mg of pioglitazone or placebo to their existing metformin and sulfonylurea therpay for four weeks in a double-blind study design. Protein risk marker levels (hsCRP, MMP-9, MCP-1, etc.) and the expression of NFκB subunits and NFκB-modulated cytokines from isolated peripheral monocyte/macrophages were determined at baseline and endpoint. There were no changes in HbA1c, but significant biomarker improvements were seen with pioglitazone only. The mRNA marker expression was downregulated by pioglitazone and further up-regulated with placebo (e.g., P105 pioglitazone: −19%/placebo: +6%, RelA: −20%/+2%, MMP−9: −36%/+9%, TNFα: −10%/+14%, P < 0.05
between groups in all cases).
Conclusions. Pioglitazone very rapidly down-regulated the activated state of peripheral monocytes/macrophages as assessed by mRNA expression of NFκB and NFκB-modulated cytokines and decreased plasma levels of cardiovascular risk marker proteins independent of glycemic control.
Obesity induced in mice by high-fat feeding activates the protein kinase cdk5 in adipose tissues. This results in phosphorylation of the nuclear receptor PPARγ, a dominant regulator of adipogenesis and fat cell gene expression, at serine 273. This modification of PPARγ does not alter its adipogenic capacity, but leads to dysregulation of a large number of genes whose expression is altered in obesity, including a reduction in the expression of the insulin-sensitizing adipokine, adiponectin. The phosphorylation of PPARγ by cdk5 is blocked by anti-diabetic PPARγ ligands, such as rosiglitazone and MRL24. This inhibition works both in vivo and in vitro, and surprisingly, is completely independent of classical receptor transcriptional agonism. Similarly, inhibition of PPARγ phosphorylation in obese patients by rosiglitazone is very tightly associated with the anti-diabetic effects of this drug. These data strongly suggest that cdk5-mediated phosphorylation of PPARγ may be involved in the pathogenesis of insulin-resistance, and present an opportunity for development of an improved generation of anti-diabetic drugs through PPARγ.
phosphorylation; PPARγ; cdk5; p35; adipogenesis; adipose tissue; rosiglitazone; MRL24
Background—Expression of pro-inflammatory
cytokines is increased in the intestinal lamina propria of patients
with inflammatory bowel disease (IBD). Nuclear factor κB (NFκB)
controls transcription of inflammation genes. On activation, NFκB is
rapidly released from its cytoplasmic inhibitor (IκB), transmigrates
into the nucleus, and binds to DNA response elements in gene promoter regions.
Aims—To investigate whether increased activation
of NFκB is important in IBD and may be down-regulated by
Methods—Activation of NFκB was determined by
western blot assessment and electrophoretic mobility shift assay in
nuclear extracts of colonic biopsy samples as well as lamina propria
Results—Nuclear levels of NFκB p65 are increased
in lamina propria biopsy specimens from patients with Crohn's disease
in comparison with patients with ulcerative colitis and controls. Increased activation of NFκB was detected in lamina propria
mononuclear cells from patients with active IBD. Corticosteroids
strongly inhibit intestinal NFκB activation in IBD in vivo and in
vitro by stabilising the cytosolic inhibitor IκBα against
activation induced degradation.
Conclusions—In both IBDs, but particularly
Crohn's disease, increased activation of NFκB may be involved in the
regulation of the inflammatory response. Inhibition of NFκB
activation may represent a mechanism by which steroids exert an
anti-inflammatory effect in IBD.
interleukin 1ß; inflammatory bowel disease; intestinal immunity; signal transduction; steroids; tumour necrosis
BACKGROUND/AIM—Proinflammatory cytokines are key factors in the pathogenesis of Crohn's disease (CD). Activation of nuclear factor kappa B (NFκB), which is involved in their gene transcription, is increased in the intestinal mucosa of CD patients. As butyrate enemas may be beneficial in treating colonic inflammation, we investigated if butyrate promotes this effect by acting on proinflammatory cytokine expression.
METHODS—Intestinal biopsy specimens, isolated lamina propria cells (LPMC), and peripheral blood mononuclear cells (PBMC) were cultured with or without butyrate for assessment of secretion of tumour necrosis factor (TNF) and mRNA levels. NFκB p65 activation was determined by immunofluorescence and gene reporter experiments. Levels of NFκB inhibitory protein (IκBα) were analysed by western blotting. The in vivo efficacy of butyrate was assessed in rats with trinitrobenzene sulphonic acid (TNBS) induced colitis.
RESULTS—Butyrate decreased TNF production and proinflammatory cytokine mRNA expression by intestinal biopsies and LPMC from CD patients. Butyrate abolished lipopolysaccharide (LPS) induced expression of cytokines by PBMC and transmigration of NFκB from the cytoplasm to the nucleus. LPS induced NFκB transcriptional activity was decreased by butyrate while IκBα levels were stable. Butyrate treatment also improved TNBS induced colitis.
CONCLUSIONS—Butyrate decreases proinflammatory cytokine expression via inhibition of NFκB activation and IκBα degradation. These anti-inflammatory properties provide a rationale for assessing butyrate in the treatment of CD.
Keywords: inflammation; butyrate; Crohn's disease; nuclear factor kappa B; cytokines
Macrophage lipid metabolism and inflammatory responses are both regulated by the nuclear receptors PPAR and LXR. Emerging links between inflammation and metabolic disease progression suggest that PPAR and LXR signaling may alter macrophage function and thereby impact systemic metabolism. In this study, the function of macrophage PPAR and LXR in Th1-biased C57BL/6 mice was tested using a bone marrow transplantation approach with PPARγ−/−, PPARδ−/−, PPARγδ−/−, and LXRαβ−/− cells. Despite their inhibitory effects on inflammatory gene expression, loss of PPARs or LXRs in macrophages did not exert major effects on obesity or glucose tolerance induced by a high-fat diet. Treatment with rosiglitazone effectively improved glucose tolerance in mice lacking macrophage PPARγ, suggesting that cell types other than macrophages are the primary mediators of the anti-diabetic effects of PPARγ agonists in our model system. C57BL/6 macrophages lacking PPARs or LXRs exhibited normal expression of most alternative activation gene markers, indicating that macrophage alternative activation is not absolutely dependent on these receptors in the C57BL/6 background under the conditions used here. These studies suggest that genetic background may be an important modifier of nuclear receptor effects in macrophages. Our results do not exclude a contribution of macrophage PPAR and LXR expression to systemic metabolism in certain contexts, but these factors do not appear to be dominant contributors to glucose tolerance in a high-fat-fed Th1-biased bone marrow transplant model.
PPAR; LXR; macrophage; inflammation; insulin resistance; immune phenotype
Several drugs approved for a variety of indications have been shown to exhibit antiangiogenic effects. Our study focuses on the PPARγ ligand rosiglitazone, a compound widely used in the treatment of type 2 diabetes. We demonstrate, for the first time to our knowledge, that PPARγ is highly expressed in tumor endothelium and is activated by rosiglitazone in cultured endothelial cells. Furthermore, we show that rosiglitazone suppresses primary tumor growth and metastasis by both direct and indirect antiangiogenic effects. Rosiglitazone inhibits bovine capillary endothelial cell but not tumor cell proliferation at low doses in vitro and decreases VEGF production by tumor cells. In our in vivo studies, rosiglitazone suppresses angiogenesis in the chick chorioallantoic membrane, in the avascular cornea, and in a variety of primary tumors. These results suggest that PPARγ ligands may be useful in treating angiogenic diseases such as cancer by inhibiting angiogenesis.
Human peritoneal mesothelial cells (MC) play an important role in inflammatory processes of the peritoneal cavity by producing various cytokines and chemokines, such as monocyte chemoattractant protein-1 (MCP-1). The present study was designed to assess the effect of the peroxisome proliferator-activated receptor-gamma- (PPARγ-) activator rosiglitazone on the mesothelial MCP-1 expression and release. Primary cultures of MC were obtained from omental tissue. MCP-1 antigen concentrations were measured in the cell supernatant by ELISA and MCP-1 mRNA levels by real-time polymerase chain reaction. The presence of PPARγ on MC was assayed in a Western Blot analysis. MC constitutively express PPARγ. Activation of this receptor via rosiglitazone (0,1–10 μmol/L) resulted in significantly reduced amounts of mesothelial MCP-1 release as well as MCP-1 mRNA. The use of the PPARγ inhibitor GW-9662 could completely prevent the rosiglitazone effects. Rosiglitazone was also effective in reducing TNFα-induced enhanced secretion of MCP-1. Our findings indicate that glitazones are effective in reducing constitutive and TNFα-stimulated mesothelial MCP-1 mRNA expression and release.
PPARγ plays a key role in adipocyte biology, and Rosiglitazone (Rosi), a thiazolidinedione (TZD)/PPARγ agonist, is a potent insulin-sensitizing agent. Recent evidences demonstrate that adipose tissue inflammation links obesity with insulin resistance and that the insulin-sensitizing effects of TZDs result, in part, from their anti-inflammatory properties. However the underlying mechanisms are unclear.
Methodology and Principal Findings
In this study, we establish a link between free fatty acids (FFAs) and PPARγ in the context of obesity-associated inflammation. We show that treatment of adipocytes with FFAs, in particular Arachidonic Acid (ARA), downregulates PPARγ protein and mRNA levels. Furthermore, we demonstrate that the downregulation of PPARγ by ARA requires the activation the of Endoplamsic Reticulum (ER) stress by the TLR4 pathway. Knockdown of adipocyte PPARγ resulted in upregulation of MCP1 gene expression and secretion, leading to enhanced macrophage chemotaxis. Rosi inhibited these effects. In a high fat feeding mouse model, we show that Rosi treatment decreases recruitment of proinflammatory macrophages to epididymal fat. This correlates with decreased chemokine and decreased chemokine receptor expression in adipocytes and macrophages, respectively.
Conclusions and Significance
In summary, we describe a novel link between FAs, the TLR4/ER stress pathway and PPARγ, and adipocyte-driven recruitment of macrophages. We thus both describe an additional potential mechanism for the anti-inflammatory and insulin-sensitizing actions of TZDs and an additional detrimental property associated with the activation of the TLR4 pathway by FA.
Inflammation plays a role in the response to metabolic stress in type 2 diabetes. However, the effects of rosiglitazone on inflammation of skeletal muscle have not been fully examined in type 2 diabetes.
We investigated the effects of the insulin-sensitizing anti-diabetic agent, rosiglitazone, on the progression of skeletal muscle inflammation in Otsuka Long-Evans Tokushima Fatty (OLETF) type 2 diabetic rats. We examined the expression of serologic markers (serum glucose, insulin and free fatty acid) and inflammatory cytokines (tumor-necrosis factor-α, interleukin [IL]-1β and IL-6) in OLETF rats from early to advanced diabetic stage (from 28 to 40 weeks of age).
Serum glucose and insulin concentrations were significantly decreased in rosiglitazone-treated OLETF rats compared to untreated OLETF rats. Rosiglitazone treatment significantly decreased the concentrations of serum inflammatory cytokines from 28 to 40 weeks of age. The mRNA expression of various cytokines in skeletal muscle was reduced in rosiglitazone-treated OLETF rats compared with untreated OLETF rats. Furthermore, rosiglitazone treatment resulted in the downregulation of ERK1/2 phosphorylation and NF-κB expression in the skeletal muscle of OLETF rats.
These results suggest that rosiglitazone may improve insulin sensitivity with its anti-inflammatory effects on skeletal muscle.
Diabetes mellitus, type 2; Inflammation; Muscle, skeletal; Rats, inbred OLETF; Rosiglitazone
Background and aims
Recent studies with μ opioid receptor (MOR) deficient mice support a physiological anti‐inflammatory effect of MOR at the colon interface. To better understand the potential pharmacological effect of certain opiates in inflammatory bowel diseases (IBD), we (1) evaluated the regulation in vivo and in vitro of human MOR expression by inflammation; and (2) tested the potential anti‐inflammatory function of a specific opiate (DALDA) in inflamed and resting human mucosa.
Patients and methods
Expression of MOR mRNA and protein was evaluated in healthy and inflamed small bowel and colonic tissues, isolated peripheral blood mononuclear cells and purified monocytes, and CD4+ and CD8+ T cells from healthy donors and IBD patients. The effect of cytokines and nuclear factor κB (NFκB) activation on MOR expression in lymphocyte T and monocytic human cell lines was assessed. Finally, DALDA induced anti‐inflammatory effect was investigated in mucosal explants from controls and IBD patients.
MOR was expressed in ileal and colonic enteric neurones as well as in immunocytes such as myeloid cells and CD4+ and CD8+ T cells. Overexpressed in active IBD mucosa, MOR was significantly enhanced by cytokines and repressed by NFκB inhibitor in myeloid and lymphocytic cell lines. Furthermore, ex vivo DALDA treatment dampened tumour necrosis factor α mRNA expression in the colon of active IBD patients.
Given the increased expression of MOR and the ex vivo beneficial effect of DALDA in active IBD, natural and/or synthetic opioid agonists could help to prevent overt pathological intestinal inflammation.
mu opioid receptors; intestinal inflammation; inflammatory bowel diseases
Type 2 diabetes has become a major burden to the health care systems worldwide. Among the drugs approved for this indication, glimepiride and rosiglitazone have gained substantial importance in routine use. While glimepiride stimulates β-cell secretion and leads to reduction of blood glucose values, rosiglitazone activates PPARγ and improves insulin resistance, at the vascular and metabolically active cells. Therefore, the combination of the two drugs may be an interesting approach to improve glycemic control and lower cardiovascular risk. A fixed combination of both drugs has been approved for clinical use in the US and EU. The combination of glimepiride and rosiglitazone is generally well tolerated and the use of a fixed combination may lead to improved adherence of the patients to their therapy. The purpose of this review is to evaluate the clinical data that have been published on this combination, appearing to represent a convenient way to obtain therapeutic targets in patients with type 2 diabetes mellitus.
rosiglitazone; glimepiride; thiazolidinediones; sulfonylurea; combination
FOXC2 has lately been implicated in diabetes and obesity as well as mitochondrial function and biogenesis and also as a regulator of mtTFA/Tfam. In this study, the expression of FOXC2 and selected genes involved in mitochondrial function and biogenesis in healthy subjects and in a matched cohort with type 2 diabetes patients before and after treatment with rosiglitazone was determined. Quantitative real time PCR was used to analyze both RNA and DNA from biopsies from subcutaneous adipose tissue.
Blood samples and subcutaneous abdominal fat biopsies were collected from 12 T2D patients, of which 11 concluded the study, pre-treatment and 90 days after initiation of rosiglitazone treatment, and from 19 healthy control subjects on the first and only visit from healthy subjects. Clinical parameters were measured on the blood samples. RNA and DNA were prepared from the fat biopsies and gene expression was measured with real time PCR.
The expression level of genes in the mitochondrial respiratory complexes I - IV were significantly downregulated in the diabetic patients and restored in response to rosiglitazone treatment. Rosiglitazone treatment also increased the relative number of mitochondria in diabetic patients compared with controls. Furthermore, the transcription factors FOXC2 and mtTFA/Tfam displayed a response pattern identical to the mitochondrial genes.
FOXC2, mtTFA/Tfam and subunits of the respiratory complexes I - IV show equivalent regulation in gene expression levels in response to TZD treatment. This, together with the knowledge that FOXC2 has a regulatory function of mtTFA/Tfam and mitochondrial biogenesis, suggests that FOXC2 has a possible functional role in the TZD activated mitochondrial response.
The nuclear accumulation and transcriptional activity of NFκB are constitutively increased in cutaneous T-cell lymphoma (CTCL) cells, and are responsible for their increased survival and proliferation. However, in addition to the anti-apoptotic and pro-inflammatory genes, NFκB induces expression of immunosuppressive genes, such as IL-10 and TGFβ, which inhibit the immune responses and are characteristic for the advanced stages of CTCL. While the mechanisms regulating NFκB-dependent transcription of anti-apoptotic and pro-inflammatory genes have been studied extensively, very little is known about the NFκB regulation of immunosuppressive genes. The specificity of NFκB-regulated responses is determined by the subunit composition of NFκB complexes recruited to the individual promoters, post-translational modifications of NFκB proteins, as well as by their interactions with other transcriptional factors and regulators. In this review, we discuss the mechanisms regulating the transcription of NFκB-dependent anti-apoptotic, pro-inflammatory and immunosuppressive genes in CTCL cells, as potential targets for CTCL therapies.
Apoptosis; bortezomib; cutaneous T cell lymphoma; IκBα; IL-10; immunosuppression; NFκB; proteasome inhibition; TGFβ
Efficient clearance of apoptotic cells (AC) by professional phagocytes is crucial for tissue homeostasis and resolution of inflammation. Macrophages respond to AC with an increase in anti-inflammatory cytokine production, but a diminished release of pro-inflammatory mediators. Mechanisms to explain attenuated pro-inflammatory cytokine formation remain elusive. We provide evidence that peroxisome proliferator-activated receptor γ (PPARγ) coordinates anti-inflammatory responses following its activation by AC. Exposing murine RAW264.7 macrophages to AC prior to LPS-stimulation, reduced NFκB transactivation and lowered target gene expression of e.g. TNFα and IL-6 compared to controls. In macrophages overexpressing a dominant negative (d/n) mutant of PPARγ, NFκB transactivation in response to LPS was restored, while macrophages from myeloid lineage-specific conditional PPARγ knockout mice proved that PPARγ transmitted an anti-inflammatory response, delivered by AC. Expressing a PPARγ-Δaa32-250 deletion mutant, we observed no inhibition of NFκB. Analyzing the PPARγ domain structures within aa32-250, we anticipated PPARγ sumoylation in mediating the anti-inflammatory effect in response to AC. Interfering with sumoylation of PPARγ by mutating the predicted sumoylation site (K77R), or knockdown of the SUMO E3 ligase PIAS1, eliminated the ability of AC to suppress NFκB. ChIP analysis demonstrated that AC prevented the LPS-induced removal of nuclear receptor co-repressor (NCoR) from the κB site within the TNFα promoter. We conclude that AC induce PPARγ sumoylation to attenuate the removal of NCoR, thereby blocking transactivation of NFκB. This contributes to an anti-inflammatory phenotype shift in macrophages responding to AC, by lowering pro-inflammatory cytokine production.
monocytes/macrophages; inflammation; phagocytosis; molecular biology
We developed KDT501, a novel substituted 1,3-cyclopentadione chemically derived from hop extracts, and evaluated it in various in vitro and in vivo models of diabetes and insulin sensitivity.
KDT501 was evaluated for anti-inflammatory effects in monocyte/macrophage cells; agonistic activity for peroxisome proliferator-activated receptors (PPAR); lipogenesis and gene expression profile in human subcutaneous adipocytes. Body composition, glucose, insulin sensitivity, and lipids were assessed in diet-induced obesity (DIO) mice and Zucker Diabetic Fatty (ZDF) rats after oral administration.
KDT501 mediated lipogenesis in 3T3L1 and human subcutaneous adipocytes; however, the gene expression profile of KDT501 differed from that of the full PPARγ agonist rosiglitazone, suggesting that KDT501 has pleiotropic biological activities. In addition, KDT501 showed only modest, partial PPARγ agonist activity and exhibited anti-inflammatory effects in monocytes/macrophages that were not observed with rosiglitazone. In a DIO mouse model, oral administration of KDT501 significantly reduced fed blood glucose, glucose/insulin AUC following an oral glucose bolus, and body fat. In ZDF rats, oral administration of KDT501 significantly reduced fed glucose, fasting plasma glucose, and glucose AUC after an oral glucose bolus. Significant, dose-dependent reductions of plasma hemoglobin A1c, weight gain, total cholesterol, and triglycerides were also observed in animals receiving KDT501.
These results indicate that KDT501 produces a unique anti-diabetic profile that is distinct in its spectrum of pharmacological effects and biological mechanism from both metformin and pioglitazone. KDT501 may thus constitute a novel therapeutic agent for the treatment of Type 2 diabetes and associated conditions.
To explore the hypothesis that CD36, a scavenger receptor and fatty acid translocase, is upregulated in peripheral blood mononuclear cells (PBMCs) among patients with type 2 diabetes and is a biomarker of PBMC activation and inflammation.
RESEARCH DESIGN AND METHODS
We used a cross-sectional observational design to study a multi-racial/ethnic population sample consisting of Caucasians, Hispanics, and Native Americans with type 2 diabetes (n = 33) and nondiabetic control subjects (n = 27). PBMC CD36 mRNA/protein and plasma high sensitivity (hs) C-reactive protein (hsCRP), hs–interleukin-6 (hsIL-6), and adiponectin were measured.
Unadjusted PBMC CD36 mRNA and protein were 1.56- and 1.63-fold higher, respectively, among type 2 diabetic subjects versus control subjects. PBMC CD36 protein was directly associated with CD36 mRNA, plasma hsCRP, and hsIL-6 and inversely associated with plasma adiponectin in both groups.
Increased CD36 expression is a biomarker of PBMC activation and inflammation and may become a useful tool in cardiovascular disease risk stratification.
The nuclear receptor peroxisome proliferator-activated receptor γ (PPARγ) is a key transcriptional regulator of both lipid metabolism and inflammation. The importance of PPARγ is accentuated by the widespread use of synthetic PPARγ agonists, thiazolidinediones (such as rosiglitazone), as drugs for insulin resistance and type II diabetes. Fractalkine (FKN) and FKN receptor (FR) play an important role in the immune responses by regulating leukocyte migration and adhesion to inflamed peripheral tissues. In this study, we have identified a novel link between PPARγ activation and FKN signaling. On one hand, the activation of PPARγ by rosiglitazone in macrophages not only represses the transcription of the FR gene, but also prevents the plasma membrane translocation of the FR protein. On the other hand, the activation of PPARγ by rosiglitazone in endothelial cells also impedes the nuclear export of FKN. Together, these data suggest that PPARγ activation represses FKN signaling. These findings indicate a previously unrecognized mechanism that may contribute to the anti-inflammatory effect of PPARγ.
At a time when the twin epidemics of obesity and type 2 diabetes threaten to engulf even the most well-resourced Western healthcare systems, the nuclear receptor peroxisome proliferator-activated receptor γ (PPARγ) has emerged as a
bona fide therapeutic target for treating human metabolic disease. The novel insulin-sensitizing antidiabetic thiazolidinediones (TZDs, e.g., rosiglitazone, pioglitazone), which are licensed for use in the treatment of type 2 diabetes, are high-affinity PPARγ ligands, whose beneficial effects extend beyond improvement in glycaemic control to include amelioration of dyslipidaemia, lowering of blood pressure, and favourable modulation of macrophage lipid handling and inflammatory responses. However, a major drawback to the clinical use of exisiting TZDs is weight gain, reflecting both enhanced adipogenesis and fluid retention, neither of which is desirable in a population that is already overweight and prone to cardiovascular disease. Accordingly, the “search is on” to identify the next generation of PPARγ modulators that will promote maximal clinical benefit by targeting specific facets of the metabolic syndrome (glucose intolerance/diabetes, dyslipidaemia, and hypertension), while simultaneously avoiding undesirable side effects of PPARγ activation (e.g., weight gain). This paper outlines the important clinical and laboratory observations made in human subjects harboring genetic variations in PPARγ that support such a therapeutic strategy.
Airway smooth muscle (ASM) cells have been reported to contribute to the inflammation of asthma. Because the thiazolidinediones (TZDs) exert anti-inflammatory effects, we examined the effects of troglitazone and rosiglitazone on the release of inflammatory moieties from cultured human ASM cells. Troglitazone dose-dependently reduced the IL-1β–induced release of IL-6 and vascular endothelial growth factor, the TNF-α–induced release of eotaxin and regulated on activation, normal T expressed and secreted (RANTES), and the IL-4–induced release of eotaxin. Rosiglitazone also inhibited the TNF-α–stimulated release of RANTES. Although TZDs are known to activate peroxisome proliferator–activated receptor-γ (PPARγ), these anti-inflammatory effects were not affected by a specific PPARγ inhibitor (GW 9662) or by the knockdown of PPARγ using short hairpin RNA. Troglitazone and rosiglitazone each caused the activation of adenosine monophosphate-activated protein kinase (AMPK), as detected by Western blotting using a phospho-AMPK antibody. The anti-inflammatory effects of TZDs were largely mimicked by the AMPK activators, 5-amino-4-imidazolecarboxamide ribose (AICAR) and metformin. However, the AMPK inhibitors, Ara A and Compound C, were not effective in preventing the anti-inflammatory effects of troglitazone or rosiglitzone, suggesting that the effects of these TZDs are likely not mediated through the activation of AMPK. These data indicate that TZDs inhibit the release of a variety of inflammatory mediators from human ASM cells, suggesting that they may be useful in the treatment of asthma, and the data also indicate that the effects of TZDs are not mediated by PPARγ or AMPK.
shRNA; anti-inflammatory; PPARγ; IL-1β; TNF-α
Type 2 diabetes often displays hyperlipidemia. We examined palmitate effects on pancreatic islet function in relation to FFA receptor GPR40, NO generation, insulin release, and the PPARγ agonistic thiazolidinedione, rosiglitazone.
Rosiglitazone suppressed acute palmitate-stimulated GPR40-transduced PI hydrolysis in HEK293 cells and insulin release from MIN6c cells and mouse islets. Culturing islets 24 h with palmitate at 5 mmol/l glucose induced β-cell iNOS expression as revealed by confocal microscopy and increased the activities of ncNOS and iNOS associated with suppression of glucose-stimulated insulin response. Rosiglitazone reversed these effects. The expression of iNOS after high-glucose culturing was unaffected by rosiglitazone. Downregulation of GPR40 by antisense treatment abrogated GPR40 expression and suppressed palmitate-induced iNOS activity and insulin release.
We conclude that, in addition to mediating acute FFA-stimulated insulin release, GPR40 is an important regulator of iNOS expression and dysfunctional insulin release during long-term exposure to FFA. The adverse effects of palmitate were counteracted by rosiglitazone at GPR40, suggesting that thiazolidinediones are beneficial for β-cell function in hyperlipidemic type 2 diabetes.
In view of the previously described anti-inflammatory effects of insulin, we investigated the potential suppressive effect of insulin on plasma concentrations and expression of the chemokines, monocyte chemoattractant protein-1 (MCP-1) and regulated on activation normal T-cell expressed and secreted (RANTES) and their receptors, chemokine receptor (CCR)-2 and CCR-5, in mononuclear cells (MNCs). We also investigated the effect of insulin on other chemokines.
RESEARCH DESIGN AND METHODS
Ten obese type 2 diabetic patients were infused with insulin (2 units/h with 100 ml of 5% dextrose/h) for 4 h. Another 8 and 6 type 2 diabetic patients were infused with 100 ml of 5% dextrose/h or saline for 4 h, respectively, and served as control subjects. Blood samples were obtained at 0, 2, 4, and 6 h.
Insulin infusion significantly suppressed the plasma concentrations of MCP-1, eotaxin, and RANTES and the expression of RANTES, macrophage inflammatory protein (MIP)-1β, CCR-2, and CCR-5 in MNCs at 2 and 4 h. Dextrose and saline infusions did not alter these indexes.
A low-dose infusion of insulin suppresses the plasma concentration of key chemokines, MCP-1, and RANTES, and the expression of their respective receptors, CCR-2 and CCR-5, in MNCs. Insulin also suppresses the expression of RANTES and MIP-1β in MNCs. These actions probably contribute to the comprehensive anti-inflammatory effect of insulin.
Obesity is associated with a chronic low-grade inflammation that predisposes to insulin resistance and the development of type 2 diabetes. In this metabolic context, gastrointestinal (GI) candidiasis is common. We recently demonstrated that the PPARγ ligand rosiglitazone promotes the clearance of Candida albicans through the activation of alternative M2 macrophage polarization. Here, we evaluated the impact of high fat diet (HFD)-induced obesity and the effect of rosiglitazone (PPARγ ligand) or WY14643 (PPARα ligand) both on the phenotypic M1/M2 polarization of peritoneal and cecal tissue macrophages and on the outcome of GI candidiasis. We demonstrated that the peritoneal macrophages and the cell types present in the cecal tissue from HF fed mice present a M2b polarization (TNF-αhigh, IL-10high, MR, Dectin-1). Interestingly, rosiglitazone induces a phenotypic M2b-to-M2a (TNF-αlow, IL-10low, MRhigh, Dectin-1high) switch of peritoneal macrophages and of the cells present in the cecal tissue. The incapacity of WY14643 to switch this polarization toward M2a state, strongly suggests the specific involvement of PPARγ in this mechanism. We showed that in insulin resistant mice, M2b polarization of macrophages present on the site of infection is associated with an increased susceptibility to GI candidiasis, whereas M2a polarization after rosiglitazone treatment favours the GI fungal elimination independently of reduced blood glucose. In conclusion, our data demonstrate a dual benefit of PPARγ ligands because they promote mucosal defence mechanisms against GI candidiasis through M2a macrophage polarization while regulating blood glucose level.
PPARγ regulates both glucose metabolism and bone mass. Recent evidence suggests that the therapeutic modulation of PPARγ activity with anti-diabetic thiazolidinediones elicits unwanted effects on bone. In this study, the effects of rosiglitazone on the skeleton of growing (1 month), adult (6 month), and aged (24 month) C57BL/6 mice were determined. Aging was identified as a confounding factor for rosiglitazone-induced bone loss that correlated with the increased expression of PPARγ in bone marrow mesenchymal stem cells. The bone of young growing mice was least affected, although a significant decrease in bone formation rate was noted. In both adult and aged animals bone volume was significantly decreased by rosiglitazone. In adult animals bone loss correlated with attenuated bone formation, whereas in aged animals bone loss was associated with increased osteoclastogenesis, mediated by increased RANKL expression. PPARγ activation led to changes in marrow structure and function such as a decrease in osteoblast number, an increase in marrow fat cells, an increase in osteoclast number, and a loss of the multipotential character of marrow mesenchymal stem cells. In conclusion, rosiglitazone induces changes in bone reminiscent of aged bone and appears to induce bone loss by altering the phenotype of marrow mesenchymal stem cells.