Browsing by Subject "Palmitoleate"

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Open Access
Double bond configuration of palmitoleate is critical for atheroprotection
(Elsevier, 2019) Çimen, I.; Yıldırım, Zehra; Doğan, Aslı Ekin; Yıldırım, Aslı Dilber; Tufanlı, Ö.; Onat, Umut İnci; Nguyen, U.; Watkins, S.; Weber, C.; Erbay, Ebru
Objective Saturated and trans fat consumption is associated with increased cardiovascular disease (CVD) risk. Current dietary guidelines recommend low fat and significantly reduced trans fat intake. Full fat dairy can worsen dyslipidemia, but recent epidemiological studies show full-fat dairy consumption may reduce diabetes and CVD risk. This dairy paradox prompted a reassessment of the dietary guidelines. The beneficial metabolic effects in dairy have been claimed for a ruminant-derived, trans fatty acid, trans-C16:1n-7 or trans-palmitoleate (trans-PAO). A close relative, cis-PAO, is produced by de novo lipogenesis and mediates inter-organ crosstalk, improving insulin-sensitivity and alleviating atherosclerosis in mice. These findings suggest trans-PAO may be a useful substitute for full fat dairy, but a metabolic function for trans-PAO has not been shown to date. Methods Using lipidomics, we directly investigated trans-PAO's impact on plasma and tissue lipid profiles in a hypercholesterolemic atherosclerosis mouse model. Furthermore, we investigated trans-PAO's impact on hyperlipidemia-induced inflammation and atherosclerosis progression in these mice. Results Oral trans-PAO supplementation led to significant incorporation of trans-PAO into major lipid species in plasma and tissues. Unlike cis-PAO, however, trans-PAO did not prevent organelle stress and inflammation in macrophages or atherosclerosis progression in mice. Conclusions A significant, inverse correlation between circulating trans-PAO levels and diabetes incidence and cardiovascular mortality has been reported. Our findings show that trans-PAO can incorporate efficiently into the same pools that its cis counterpart is known to incorporate into. However, we found trans-PAO's anti-inflammatory and anti-atherosclerotic effects are muted due to its different structure from cis-PAO.
Open Access
Lipotoxic endoplasmic reticulum stress-associated inflammation : molecular mechanisms and modification by a bioactive lipokine
(2012) Demirsoy, Şeyma
Physiologic or pathologic processes that disturb protein folding in the endoplasmic reticulum (ER) activate a signaling pathway named the unfolded protein response (UPR). UPR promotes cell survival by reducing misfolded protein levels. The three proximal stress sensors of the UPR are known as PKR-resemble like ER kinase (PERK), inositol-requiring enzyme-1 (IRE1) and activating transcription factor 6 (ATF6), which monitor the quality of protein folding in the ER membrane and relay that information to the rest of the cell. If ER homeostasis can not be restored, prolonged UPR signaling can lead to cell death. Recent studies have shown metabolic overload, particularly high levels of fatty acids and cholesterol can induce ER stress and activate UPR signaling. These studies also demonstrated ER stress is a central mechanism that underlies the pathogenesis of metabolic diseases including obesity, type 2 diabetes, insulin resistance, atherosclerosis and hepatosteatosis. Understanding how nutrient excess activates the UPR and its novel molecular mechanisms of operation during metabolic stress could facilitate the development of novel and effective future therapeutics aiming to restore ER homeostasis. The molecular mechanisms of lipid induced activation of UPR and how the three proximal UPR stress sensors are linked to lipid metabolism and inflammation is not well understood. One of the UPR stress sensors, PERK, is a trans-membrane serine/threonine kinase with only two known downstream substrates, the eukaryotic translation initiation factor (eIF2) that controls translation initiation, and an antioxidant transcription factor, Nuclear factor eryhthroid-2-related factor-2 (Nrf2), that keeps redox homeostasis. One of the existing road blocks in studying PERK signaling has been the lack of molecular or chemical tools to regulate its activity. For my thesis studies, I developed a chemical-genetic approach to specifically modify PERK’s kinase activity. In this approach, the ATP binding pocket of a particular kinase is altered via site-directed mutagenesis in order to accommodate a bulky ATP analog that is not an effective substrate for the wild type kinase. Thus, only the mutated kinase can be targeted by the activatory or inhibitory bulky ATP analogs and this form of the kinase is referred to as ATP analog sensitive kinase (ASKA). Furthermore, I identified specific siRNA sequences that can be efficiently delivered to mouse macrophages and significantly reduce PERK expression. Both of these methods can be applied to study the direct impact of PERK activity on lipotoxic ER stress- associated inflammation. The results of the siRNA mediated PERK expression silencing experiments showed that PERK has a direct contribution to lipid-induced pro-inflammatory response in macrophages. Finally, I examined whether palmitoleate, a bioactive monounsaturated fatty acid previously shown to reduce lipid-induced ER stress and death, could also modify lipotoxic ER stress-associated inflammation. Based on the results from my experiments, palmitoleate is highly effective in preventing lipid induce inflammation. Unexpectedly, I also observed that palmitoleate could significantly block LPS-induced inflammation, too. In summary, during my thesis study I generated several useful tools including siRNA mediated knock-down of PERK and a novel chemical-genetic tool to directly and specifically modify PERK kinase activity. The findings from my studies demonstrate that PERK plays a significant role in lipid-induced inflammation, suggesting modification of PERK activity or its direct pro-inflammatory substrates could become desirable approaches to inhibit obesity-induced inflammation that contributes to the pathogenesis of diabetes and atherosclerosis. The outcome of my studies also showed that palmitoleate can significantly reduce lipotoxic-ER stress associated inflammation, which may explain its beneficial impact on both insulin resistance and atherosclerosis. Furthermore, the ATP-analog sensitive PERK mutant developed in my thesis can be coupled with proteomics to identify the full repertoire of PERK substrates during metabolic stress. In conclusion, the findings and tools developed in my thesis studies can form the basis of future studies to identify the molecular details of PERK’s involvement in lipid induced inflammation, the identification of novel PERK substrates during metabolic stress and the development of new therapeutic strategies against metabolically induced inflammation in obesity, diabetes and atherosclerosis.
Open Access
Prevention of fatty acid-induced inflammasome activation by a bioactive lipokine
(2014-08) Koyuncu, Seda
Exposure to excess lipids such as fatty acids and cholesterol leads to cellular stress, release of reactive oxygen species (ROS), inflammation dysfunction and death. These responses have important role in the pathogenesis of chronic metabolic and inflammatory diseases such asobesity, diabetes and atherosclerosis. Recent studies show that fatty acids also cause the formation of inflammatoryprotein complexes that are called inflammasome. Inflammasome promotesthe activation of caspase-1 protein and cleavage of inactive interleukin-1 beta(IL-1β) and interleukin-18 (IL-18) into their active, secreted forms.Two signals are required for the activation of inflammasome. The first signal (also known as priming step) is needed for the inducing the expression ofthe proinflammatory factors pro-IL-1β and pro-IL-18 andthe nucleotide-binding oligomerization domain receptor (NOD-like receptor) family, pyrin domain containing-3 (NLRP3) proteins through the activation of nuclear factor kappa beta (NF-κB),a transcription factor. The second signal is needed for the activation of caspase-1 and formation of the inflammasome complex. Saturated free fatty acids such as palmitate are known to cause the activation of inflammasome through generation of mitochondrial ROS as a second signal for inflammasome complex formation. In this study, I investigated the effect of palmitoleate, a bioactive monounsaturated fatty acidpreviously shown to counteract lipid-induced ER stress and enhance insulin sensitivity, on palmitate-induced activation of inflammasome complex. I observed that palmitoleate lead to a significant reduction in palmitate-stimulatedIL-1β transcription. Moreover, it reduced the expression of palmitate-stimulated secondary proinflammatory factors such as tumor necrosis factor alpha(TNF-a).Palmitoleate also diminished the palmitate induced activation of caspase-1, maturation and secretion of IL-1β in macrophages. To further understand the mechanism of the protective role of palmitoleate on palmitate induced inflammasome activation,I analyzed the effect of palmitoleate on palmitate-induced mtROS in macrophages cells were analyzed. These studies showed palmitoleate decreased palmitate induced mitochondrial oxygen species in macrophages. Moreover, I investigated the effect of palmitoleate on palmitate-induced inactivation of 5' AMP-activated protein kinase (AMPK)controls autopaghy and clearance of dysfunctional mitochondria, thereby reducing mtROS formation. Palmitoleate blocked the suppression of AMPK phosphorylation that was suppressed by palmitate, suggesting PAO's impact on mtROS secretion and inflammasome activation occurs by this upstream mechanism. Saturated fatty acids like palmitate are also known to cause endoplasmic reticulum (ER) stress, whichbe counteracted by palmitolate treatment. When ER stress happens, cells try to solve stress by activating the unfolded protein response (UPR). There are three arms of the UPR are Protein Kinase R-resemble like protein kinase (PERK), inositol-requiring enzyme-1 (IRE1) and activating factor 6 (ATF6). In this thesis, contribution of PERK and IRE1 on palmitate-induced activation of inflammasome was investigated. First, the relation between PERK and IRE1 with palmitate-stimulated mtROS was analyzedusing PERK- and IRE1- deficient mouse embryonic fibroblast cells (MEFs).The results of this study show PERK leads to a marked reduction in the formation of mtROS while IRE1 enhanced palmitate induced mtROS.Furthermore, palmitoleate suppressed palmitate-induced autophosphorylation of IRE1 while there was no effect of palmitoleate on palmitate-induced phosphorylation of PERK. In summary, palmitate-induced inflammasome activation can be inhibited by palmitoleate in boththe priming step and the second step.Palmitoleate blocks palmitate-induced suppression of phosphorylation of AMPK, leading to its reactivation and subsequent reduction in mtROS formation. Palmitate is a well-known inducer of ER stress, which can be counteracted by palmitoleic acid. Moreover, the IRE1 branch of the UPR has been shown to control inflammasome activationby regulating of mtROS production under lipotoxicity. The outcome of my studies show that the UPR branches initiated by PERK and IRE1 regulate mtROS production and therefore, palmitoleate may also block inflammasome activation induced by palmitate through this alternative mechanism.These findings imply that palmitoleate may have therapeutic applications inthe management of diseases where inflammasome activation has been shown to play a causal role such as obesity, diabetes and atherosclerosis.