Fetuin-A regulates adipose tissue macrophage content and activation in insulin resistant mice through MCP-1 and iNOS: involvement of IFNγ-JAK2-STAT1 pathway.

In the context of obesity-induced adipose tissue (AT) inflammation, migration of macrophages and their polarization from predominantly anti-inflammatory to proinflammatory subtype is considered a pivotal event in the loss of adipose insulin sensitivity. Two major chemoattractants, monocyte chemoattractant protein-1 (MCP-1) and Fetuin-A (FetA), have been reported to stimulate macrophage migration into inflamed AT instigating inflammation. Moreover, FetA could notably modulate macrophage polarization, yet the mechanism(s) is unknown. The present study was undertaken to elucidate the mechanistic pathway involved in the actions of FetA and MCP-1 in obese AT. We found that FetA knockdown in high fat diet (HFD) fed mice could significantly subdue the augmented MCP-1 expression and reduce adipose tissue macrophage (ATM) content thereby indicating that MCP-1 is being regulated by FetA. Additionally, knockdown of FetA in HFD mice impeded the expression of inducible nitric oxide synthase (iNOS) reverting macrophage activation from mostly proinflammatory to anti-inflammatory state. It was observed that the stimulating effect of FetA on MCP-1 and iNOS was mediated through interferon γ (IFNγ) induced activation of JAK2-STAT1-NOX4 pathway. Furthermore, we detected that the enhanced IFNγ expression was accounted by the stimulatory effect of FetA upon the activities of both cJun and JNK. Taken together, our findings revealed that obesity-induced FetA acts as a master upstream regulator of AT inflammation by regulating MCP-1 and iNOS expression through JNK-cJun-IFNγ-JAK2-STAT1 signaling pathway. This study opened a new horizon in understanding the regulation of ATM content and activation in conditions of obesity-induced insulin resistance.

Attenuation of macrophage accumulation and polarisation in obese diabetic mice by a small molecule significantly improved insulin sensitivity.

Accumulation and polarization of anti-inflammatory M2 to proinflammatory M1 macrophage in the adipose tissue of obese diabetic mice is an important pathogenic signature. It worsens lipid induced inflammation and insulin resistance. Here we demonstrate that a small molecule, a peroxyvanadate compound i.e. DmpzH [VO(O2)2 (dmpz)] or dmp, could robustly decrease macrophage infiltration, accumulation and their polarization in high fat diet (HFD) induced obese diabetic mice. In searching the underlying mechanism it was revealed that SIRT1 level was strikingly low in the inflamed adipose tissue of HFD mice as compared to mice fed with standard diet (SD). Administration of dmp markedly increased SIRT1 level by inducing its gene expression with a consequent decrease in macrophage population. Elevation of SIRT1 coincided with the decrease of MCP1, Fetuin-A (FetA) and IFNγ. Since MCP1 and FetA drive macrophage to inflamed adipose tissue and IFNγ promotes M2 to M1 transformation, both recruitment and M1 induced inflammation were found to be significantly repressed by dmp. In addressing the question about how dmp induced excess SIRT1 could reduce MCP1, FetA and IFNγ levels, we found that it was due to the inactivation of NFκB because of its deacetylation by SIRT1. Since NFκB is the transcriptional regulator of these molecules, their expressions were significantly suppressed and that caused sharp decline in macrophage recruitment and their polarity to M1. This effected a marked fall in proinflammatory cytokine level which significantly improved insulin sensitivity. dmp is likely to be the first molecule that rescues inflammatory burden contributed by macrophage in obese diabetic mice adipose tissue which causes significant increase in insulin sensitivity therefore it may be a meaningful choice to treat type 2 diabetes.

Nrf2 inhibition regulates intracellular lipid accumulation in mouse insulinoma cells and improves insulin secretory function.

High amount of fat in the pancreas is linked to poor functioning of ß-cells and raises the risk of type 2 diabetes. Here we report the putative role of a circulatory glycoprotein Fetuin-A, a known obesity marker, in promoting lipid accumulation in ß-cells and its association with Fatty acid translocase/CD36 for lipid storage culminate in ß-cell dysfunction. Additionally, this work reveals regulation of CD36 via Nrf2, a key regulator of oxidative stress, and reduction of lipid accumulation by suppression of Nrf2 that restores ß-cell function. Palmitate (0.50 mM) and Fetuin-A (100 µg/mL) exposure showed high levels of intracellular lipid in MIN6 (mouse insulinoma cells) with a concomitant decrease in insulin secretion. This also increased the expression of important lipogenic factors, like CD36, PGC1α, PPARγ, and SREBP1. Flow cytometry analysis of CD36 membrane localization has been corroborated with an increased accumulation of lipids as indicated by Oil-Red-O staining. Immunoblotting and immunofluorescence of Nrf2 indicated its high expression in palmitate-fetuin-A incubation and translocation in the nucleus. Suppression of Nrf2 by siRNA showed a reduced expression of lipogenic genes, ablation of lipid droplets, decrease in the number of apoptotic cells, and restoration of insulin secretion with a corresponding increase of Pdx1, BETA2, and Ins1 gene expression. Our study thus suggested an important aspect of lipid accumulation in the pancreatic ß-cells contributing to ß-cell dysfunction and demonstrated the role of Fetuin-A in CD36 expression, with a possible way of restoring ß-cell function by targeting Nrf2.

Adipose tissue macrophages and their role in obesity-associated insulin resistance: an overview of the complex dynamics at play.

Obesity, a major global health concern, is characterized by serious imbalance between energy intake and expenditure leading to excess accumulation of fat in adipose tissue (AT). A state of chronic low-grade AT inflammation is prevalent during obesity. The adipose tissue macrophages (ATM) with astounding heterogeneity and complex regulation play a decisive role in mediating obesity-induced insulin resistance. Adipose-derived macrophages were broadly classified as proinflammatory M1 and anti-inflammatory M2 subtypes but recent reports have proclaimed several novel and intermediate profiles, which are crucial in understanding the dynamics of macrophage phenotypes during development of obesity. Lipid-laden hypertrophic adipocytes release various chemotactic signals that aggravate macrophage infiltration into AT skewing toward mostly proinflammatory status. The ratio of M1-like to M2-like macrophages is increased substantially resulting in copious secretion of proinflammatory mediators such as TNFα, IL-6, IL-1ß, MCP-1, fetuin-A (FetA), etc. further worsening insulin resistance. Several AT-derived factors could influence ATM content and activation. Apart from being detrimental, ATM exerts beneficial effects during obesity. Recent studies have highlighted the prime role of AT-resident macrophage subpopulations in not only effective clearance of excess fat and dying adipocytes but also in controlling vascular integrity, adipocyte secretions, and fibrosis within obese AT. The role of ATM subpopulations as friend or foe is determined by an intricate interplay of such factors arising within hyperlipidemic microenvironment of obese AT. The present review article highlights some of the key research advances in ATM function and regulation, and appreciates the complex dynamics of ATM in the pathophysiologic scenario of obesity-associated insulin resistance.

Increase in PPARγ inhibitory phosphorylation by Fetuin-A through the activation of Ras-MEK-ERK pathway causes insulin resistance.

Obesity induced insulin resistance is primarily regulated by the inhibitory phosphorylation of peroxisome proliferator-activated receptor γ at serine 273 (PPARγS273) which has been shown to be regulated by MEK and ERK. An upstream regulatory molecule of this pathway could be a therapeutic option. Here we analyzed the involvement of Fetuin-A (FetA), a key hepato-adipokine implicated in insulin resistance, as an upstream regulator molecule for the regulation of PPARγ inhibitory phosphorylation. Mice fed with standard diet (SD), high fat diet (HFD) and HFD with FetA knockdown (HFD-FetAKD) were used to examine the role of FetA on PPARγS273 phosphorylation in adipocytes. The mechanism of regulation and its effect on skeletal muscle were studied using primary adipocytes, 3T3-L1 (preadipocyte) and C2C12 (myotube) cell lines. Increased FetA in HFD mice strongly correlated with augmentation of PPARγS273 phosphorylation in inflamed adipocytes while knockdown of FetA suppressed it. This effect of FetA was mediated through the activation of Ras which in turn activated MEK and ERK. On addressing how FetA could stimulate activation of Ras, we found that FetA triggered TNFα in inflamed adipocytes which induced Ras activation. The ensuing sharp fall in adiponectin level attenuated AMPK activation in skeletal muscle cells affecting mitochondrial ATP production. Our data reveal the essential role of FetA induced activation of Ras in regulating PPARγ inhibitory phosphorylation through Ras-MEK-ERK pathway which downregulates adiponectin disrupting skeletal muscle mitochondrial bioenergetics. Thus, FetA mediated PPARγ inactivation has adverse consequences upon adipocyte-myocyte crosstalk leading to disruption of energy homeostasis and loss of insulin sensitivity.

Impairment of energy sensors, SIRT1 and AMPK, in lipid induced inflamed adipocyte is regulated by Fetuin A.

Although several reports demonstrated that accumulation of excess lipid in adipose tissue produces defects in adipocyte which leads to the disruption of energy homeostasis causing severe metabolic problems, underlying mechanism of this event remains yet unclear. Here we demonstrate that FetuinA (FetA) plays a critical role in the impairment of two metabolic sensors, SIRT1 and AMPK, in inflamed adipocytes of high fat diet (HFD) mice. A linear increase in adipocyte hypertrophy from 10 to 16 week was in tandem with the increase in FetA and that coincided with SIRT1 cleavage and decrease in pAMPK which adversely affects PGC1α activation. Knock down (KD) of FetA gene in HFD mice could significantly improve this situation indicating FetA's contribution in the damage of energy sensors in inflamed adipocyte. However, FetA effect was not direct, it was mediated through TNF-α which again is dependent on FetA as FetA augments TNF-α expression. FetA being an upstream regulator of TNF-α, its suppression prevented TNF-α mediated Caspase-1 activation and cleavage of SIRT1. FetA induced inactivation of PGC1α due to SIRT1 cleavage decreased PPARϒ, adiponectin, NRF1 and Tfam expression. All these together caused a significant fall in mitochondrial biogenesis and bioenergetics that disrupted energy homeostasis resulting loss of insulin sensitivity. Taken together, our findings revealed a new dimension of FetA, it not only induced inflammation in adipocyte but also acts as an upstream regulator of SIRT1 cleavage and AMPK activation. Intervention of FetA may be worthwhile to prevent metabolic imbalance that causes insulin resistance and type 2 diabetes.