Endoplasmic reticulum stress is involved in the connection between inflammation and autophagy in type 2 diabetes.

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Contents lists available at ScienceDirect

General and Comparative Endocrinology journal homepage: www.elsevier.com/locate/ygcen

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Review

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Endoplasmic reticulum stress is involved in the connection between inflammation and autophagy in type 2 diabetes

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Han Liu, Ming-ming Cao, Yang Wang, Le-chen Li, Li-bo Zhu, Guang-ying Xie, Yan-bo Li ⇑ Department of Endocrinology, The First Affiliated Hospital of Harbin Medical University, No. 23 You zheng Street Nan Gang District, Harbin 150001, China

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Article history: Received 31 May 2014 Revised 30 August 2014 Accepted 16 September 2014 Available online xxxx Keywords: Type 2 diabetes Autophagy Inflammation Endoplasmic reticulum stress Interleukin 1b

a b s t r a c t Type 2 diabetes is a chronic inflammatory disease. A number of studies have clearly demonstrated that cytokines such as interleukin 1b (IL1b) contribute to pancreatic inflammation, leading to impaired glucose homeostasis and diabetic disease. There are findings which suggest that islet b-cells can secrete cytokines and cause inflammatory responses. In this process, thioredoxin-interacting protein (TXNIP) is induced by endoplasmic reticulum (ER) stress, which further demonstrates a potential role for ER stress in innate immunity via activation of the NOD-like receptor (NLRP) 3/caspase1 inflammasome and in diabetes pathogenesis via the release of cytokines. Recent developments have also revealed a crucial role for the autophagy pathway during ER stress and inflammation. Autophagy is an intracellular catabolic system that not only plays a crucial role in maintaining the normal islet architecture and intracellular insulin content but also represents a form of programmed cell death. In this review, we focus on the roles of autophagy, inflammation, and ER stress in type 2 diabetes but, above all, on the connections among these factors. Ó 2014 Elsevier Inc. Published by Elsevier Inc. All rights reserved.

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1. Introduction Inflammation has been implicated in the pathophysiology of type 2 diabetes. However, unlike type 1 diabetes, the islet inflammatory response of type 2 diabetes is ‘‘low grade’’ and its role in the pathophysiology of type 2 diabetes is somewhat controversial (Imai et al., 2013). It has been shown that elevated levels of IL1b, IL6, MCP1, and C-reactive protein are predictive of type 2 diabetes (Donath and Shoelson, 2011). Further evidence supports the idea that overnutrition and insulin resistance result in the production of proinflammatory cytokines, such as IL1b, and the activation of signaling pathways that cause pancreatic b-cell death and dysfunction (Wang et al., 2013). Moreover, other research has shown that

Abbreviations: IL1b, interleukin 1b; TXNIP, thioredoxin-interacting protein; ER, endoplasmic reticulum; NLRP, NOD-like receptor; PERK, protein kinase RNA-like ER kinase; IRE, inositol requiring enzyme; Atg7, autophagy-related gene; LC3, light chain 3; eIF2a, eukaryotic translation initiation factor 2a; TNFa, tumor necrosis factor alpha; IFNc, interferonc; IKKB, inhibitors of NF-kB, kinase B; NF-kB, nuclear factor-KB; JNK, c-Jun N-terminal kinase; ROS, reactive oxygen species; ATF, activating transcription factor; ASC, apoptosis-associated speck-like protein containing a CARD; PA, palmitate; 3-MA, 3-methyadenine; Pdx1, pancreas duodenal homeobox 1; ULK1, UNC51-like kinase 1; PE, phosphatidylethanolamine; Atg7Db-cell, pancreaticb-cell specific Atg7-knockout. ⇑ Corresponding author. Fax: +86 451 85555637. E-mail address: [email protected] (Y.-b. Li).

ER stress activates IL1b production by the NLRP3 inflammasome through the protein kinase RNA-like ER kinase (PERK) and inositol requiring enzyme (IRE)1 pathways and mediates b-cell death (Oslowski et al., 2012). Additionally, an emerging body of evidence identifies autophagy as a critical modulator of the two major pathological arms of type 2 diabetes–impaired insulin secretion and insulin sensitivity. Autophagy is a cellular process that not only degrades proteins but also breaks down lipids, DNA and RNA. In this way, autophagy provides new pools of raw material for anabolic processes and drives a continuous flow of materials in a degradation–regeneration cycle within the cell (Rabinowitz and White, 2010). Autophagy may also contribute to programmed cell death, called autophagic death, which is different from apoptosis, depending on the cellular and environmental context (Tsujimoto and Shimizu, 2005). Thus, it is difficult to say whether autophagy promotes cell death or protects cells from diverse types of injuries, depending on the cellular and environmental context. Recent studies have reported that autophagy-related gene (Atg7) induced excessive autophagic activation in pancreatic INS-1(823/13) cells exposed to saturated fatty acids. Cathepsin B, which is induced by Atg7, contributed to the Atg7-induced NLRP3-dependent proinflammatory response and resulted in aggravation of lipotoxicity independent of apoptosis in the INS-1(823/13) cell line (Li et al., 2013). In addition, Atg16L1 is an essential component of the autophagic machinery responsible

http://dx.doi.org/10.1016/j.ygcen.2014.09.006 0016-6480/Ó 2014 Elsevier Inc. Published by Elsevier Inc. All rights reserved.

Please cite this article in press as: Liu, H., et al. Endoplasmic reticulum stress is involved in the connection between inflammation and autophagy in type 2 diabetes. Gen. Comp. Endocrinol. (2014), http://dx.doi.org/10.1016/j.ygcen.2014.09.006

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for regulation of endotoxin-induced IL1b production (Saitoh et al., 2008). ER stress also plays an active role in the process of autophagyassociated management in type 2 diabetes (Bartolome et al., 2012). Although the molecular mechanism remains unclear, microtubuleassociated protein 1 light chain 3 (LC3) can be converted to lipidated LC3 (LC3-II), and this process is reportedly mediated by PERK/ eukaryotic translation initiation factor 2a (eIF2a) phosphorylation (Gonzalez et al., 2011). Based on the above analysis, we deduce that autophagy, inflammation and ER stress are crucial in type 2 diabetes and closely connected with each other. Thus, understanding the mechanisms of their involvement in the regulation of diabetic disease may help to identify novel therapeutic targets with important clinical applications.

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2. Type 2 diabetes: a chronic inflammatory disease

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Type 2 diabetes is traditionally characterized by insulin resistance/reduced systemic insulin sensitivity and islet b-cell dysfunction. The following three mechanisms in particular are associated with insulin resistance following b-cell depletion and, indirectly, immune attack: lipotoxicity, glucotoxicity, and inflammation (Odegaard and Chawla, 2012). Recently, a number of studies have distinctly demonstrated that chronic tissue inflammation is a key contributing factor to type 2 diabetes. Elevated levels of glucose and lipids, particularly saturated fatty acids, are characteristics of insulin resistance and synergize within the b-cell to drive parallel increases in FAS expression (Unger, 1995). This contributes to the pathogenesis of type 2 diabetes via ER stress and the generation of reactive oxygen species, both of which culminate in inflammatory cytokine secretion (Harding and Ron, 2002; Hotamisligil, 2010). In particular, IL1b secretion has been known to be a mediator of b-cell dysfunction and death for more than 25 years, and its effects are potentiated by tumor necrosis factor alpha (TNFa); interferonc (IFNc) (Mandrup-Poulsen et al., 1985; Pukel et al., 1988; Hotamisligil, 2010). Additionally, a recent study indicated that IL6 and IL10 are important physiological contributors to the central insulin and leptin actions mediated by exercise, thus linking to hypothalamic ER stress and inflammation. The impairment of hypothalamic insulin and leptin signaling pathways is sufficient to promote hyperphagia, obesity, and T2D. This study also showed that the activation of inhibitors of NF-kB, kinase B (IKKB)/nuclear factor-KB (NF-kB) through elevated endoplasmic reticulum stress in the hypothalamus is associated with central insulin resistance. In this process, examination of ER stress markers demonstrated increased levels of PERK phosphorylation and of c-Jun N-terminal kinase (JNK) and IKKB activity (Ropelle et al., 2010). The most noteworthy feature of type 2 diabetes, unlike type 1 diabetes, is that the islet inflammatory response is ‘‘low-grade’’ and its role in the pathophysiology of type 2 diabetes is somewhat controversial. The latest research shows NKp46 is involved in the killing of murine b-cells in type 1 diabetes (Imai et al., 2013). However, a totally different argument could be made in type 2 diabetes. Insulin depletion in diabetic b-cells may protect them from NK cell attack and thus should be viewed as a protective response, thereby preventing NKp46-mediated b-cell destruction during low-grade inflammation (Gur et al., 2013).

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3. ER stress is involved in inflammation in the pathogenesis of type 2 diabetes A chronic excess of metabolic factors (such as lipids, glucose and cytokines), intracellular calcium and free radicals [such as reactive oxygen species (ROS) and nitric oxide] can trigger not only inflammation but also ER stress, which further disrupts metabolic

functions and thereby causes more inflammation (Zhang and Kaufman, 2008). However, this cycle of inflammation is transient and NF-kB-dependent (Böni-Schnetzler et al., 2008). Under such conditions, failure of the ER’s adaptive capacity results in activation of the UPR (Hotamisligil, 2010). Moreover, during ER stress, excess accumulation of unfolded or misfolded proteins causes cells to consume extra reduced glutathione to correctly fold these aberrantly assembled proteins, thus adding to cellular stress. Consequently, ER stress can lead to oxidative stress, which might trigger an inflammatory state, as discussed earlier (Alfadda and Sallam, 2012). Recently, Oslowski et al. showed that TXNIP (an early response gene highly induced by diabetes and hyperglycemia, encoding an endogenous inhibitor of the antioxidant thioredoxin) is a critical signaling node that links ER stress and inflammation. TXNIP has a demonstrated potential role in innate immunity during diabetes via activation of the NLRP3 inflammasome and release of IL1b, which mediates oxidative stress and ER-stress-mediated b-cell death (Zhou et al., 2010; De Nardo and Latz, 2011; Oslowski et al., 2012). These data have demonstrated that the transcriptional expression of TXNIP is regulated by ChREBP and activating transcription factor (ATF) 5. It has been shown that ChREBP expression was induced by ER stress and significantly decreased in PERK knockdown INS-1 832/13 cells and PERK knockout mouse embryonic fibroblasts. Additionally, it has been shown that PERK-mediated eIF2a phosphorylation directs the protein translation and mRNA transcription of ATF5 and that ATF5 is integral to the eIF2a kinase response. In Ire1a/PERK knockdown INS-1 832/ 13 cells and MEFs, TXNIP expression was modestly attenuated compared to control cells under ER stress conditions. In conclusion, the expression of TXNIP induced by ER stress is under the control of the IRE1a and PERK-eIF2a pathways of the UPR. This experiment proved that IL1b and IL6 upregulation was attenuated in TXNIP knockdown cells compared to control cells (Oslowski et al., 2012). Moreover, TXNIP binds to and inhibits thioredoxin and thereby can modulate the cellular redox state and promote oxidative stress (Patwari et al., 2006). These findings support the conclusion that a variety of stress signaling pathways converge at TXNIP and lead to inflammasome activation and IL1b production. IL1b is usually involved in the activation of a protein complex termed the inflammasome. The NLRP subfamily (NLRP1, NLRP3 and NLRC4), and the PYHIN family protein absence in melanoma 2 have been shown to form inflammasomes, which are high molecular weight signaling platforms. Among these is NLRP3, which contains the adaptor molecule apoptosis-associated speck-like protein containing a CARD (ASC) and pro-caspase1. Activation induces oligomerization of the NLRP3 inflammasome and recruits ASC through homotypic PYD–PYD interaction. Then, ASC recruits procaspase1 leading to autocatalytic activation of caspase1, and the active caspase1 hetero-tetramers are able to convert inactive pro-IL1b and pro-IL18 into their bioactive and secreted forms (De Nardo and Latz, 2011). Patterns or danger-associated molecular patterns activate the inflammasome. Once IL1b leaves the cell, it binds to the IL1 receptor and causes inflammation. Both potassium (K+) efflux and an increase in ROS are necessary for the activation of the NLRP3 inflammasome in response to all stimuli tested thus far. Recent studies suggest that ER stress, like other NLRP3 activators, activates the NLRP3 inflammasome in a K+ efflux- and ROS-dependent manner that may also affect the mitochondria (Menu et al., 2012). Hence, we infer that ER stress initiates a signal that is transmitted to mitochondria and then relayed to the NLRP3 inflammasome. Inflammatory cytokines converge on IKKB (an inhibitor of the kappa light polypeptide gene enhancer in b-cells), and mitogenactivated protein kinase 8/JNK1 to directly inhibit insulin action via serine phosphorylation of insulin receptor substrate 1 and 2


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(Odegaard and Chawla, 2012). b-cell damage and concomitant innate immune activation within the islet cells initiates b-cellspecific cytotoxic T lymphocyte responses, which further damage the beleaguered b-cells (Coppieters et al., 2012). Taken together, not only the expression of TXNIP but also the activation of the NLRP3 inflammasome is associated with ER stress. Therefore, in this review, we highlight the areas of ER stress and the potential effects in diabetes.

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4. Autophagy in type 2 diabetes

(Liang et al., 1999). Reducing Becn1 expression in Pdx1-deficient MIN6 cells led to an increase in the cleavage of caspase3, and this was associated with a substantial increase in cell death, which demonstrates that a minimum basal level of autophagy is required for MIN6 cell survival (Fujimoto et al., 2009). Although the study indicated that enhanced autophagy can result in deterioration of b-cell function, it is not known whether autophagy contributes to pancreatic b-cell death and reduced b cell mass, and does not necessarily mean that enhanced autophagy is harmful in patients with type 2 diabetes.

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4.1. The dual nature of autophagy in type 2 diabetes

4.2. Molecular mechanisms of autophagy

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In nutrient-rich conditions, mammalian target of rapamycin complex 1 kinase is incorporated into the UNC51-like kinase 1 (ULK1)-Atg 13-FIP200 complex, which is required during autophagy, and phosphorylates ULK1. Dephosphorylated ULK1 is enzymatically active and phosphorylates Atg13 and FIP200 to initiate the autophagic process (Mizushima, 2010). Following autophagy induction, Beclin 1 (Bcl-2-interacting protein) is liberated from Bcl-2 and forms complexes with Vps34, Vps15, and Atg14L to induce autophagosome formation or with Vps34, Vps15, and UVRAG to induce autophagosome maturation (Matsunaga et al., 2009). However, recent data demonstrate its involvement in the proper localization of another crucial autophagy-inducing complex, the phosphatidylinositol-3-kinase class3 complex (PIK3C3, p150, Ambra1 and Beclin 1 form the core of the PIK3C3 complex), which can further bind either UVRAG or Atg14L (He and Levine, 2010). Within this complex, Beclin-1 (Atg6) constitutes a platform for the binding of several interactors regulating the kinase activity of PIK3C3 and generates PI3P (phosphatidyl-inositol-3-phosphate) (Funderburk et al., 2010). Then, PI3P recruits additional double FYVE-containing protein 1 and Atg proteins to the site of autophagosome cradle formation (Quan and Lee, 2013). In spite of the aforementioned studies, further studies of autophagosome biogenesis are needed. Two ubiquitin-like conjugation systems are implicated in autophagosome membrane expansion and shaping; furthermore, the Atg system is indispensable for autophagosome completion and is similar to the ubiquitination system. In the first ubiquitin-like conjugation pathway, Atg12, Atg7, Atg10, Atg5, and Atg16L1 participate and form a large multimeric complex. The second ubiquitin-like conjugation pathway involves the phosphorylation of microtubule-associated LC3, which is finally conjugated to phosphatidyl ethanolamine (PE) to produce LC3-PE (also called LC3-II). The Atg4, Atg7, Atg3, and Atg16L1 complex also takes part in this process (Wirawan et al., 2012). The physiological role of autophagy in pancreatic b-cells has been studied, and it has been demonstrated that Atg7 is a key enzyme involved in Atg12–Atg5 conjugation and LC3 lipidation, both of which are essential for isolation membrane elongation (Ebato et al., 2008; Wirawan et al., 2012). A pancreatic b-cell specific Atg7-knockout (Atg7Db-cell) mouse model resulted in degradation of islets and impaired glucose tolerance. These mice developed hyperglycemia but not diabetes, suggesting that ‘‘basal autophagy’’ is important for the maintenance of normal islet architecture and function (Ebato et al., 2008). Therefore, expression of Atg7 is considered essential for autophagy function. Nevertheless, little is known thus far concerning the molecular mechanisms by which b-cells cope with various stresses and cellular damage associated with the reduction of insulin secretion. This may be accelerated by increased insulin resistance and needs further study.

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4.3. Autophagy and insulin resistance

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Insulin resistance in sensitive target tissues is one risk factor for type 2 diabetes. In the preclinical period of the disease, pancreatic

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The effect of autophagy in diabetes is two-sided. Autophagy is a cellular process that sequesters senescent or damaged organelles/ proteins in autophagosomes and allows the recycling of their broken down products (Levine and Kroemer, 2008). In addition, autophagy serves as a crucial element of stress responses to protect pancreatic b-cells under insulin-resistant conditions (Fujitani et al., 2009). It has also been indicated that autophagy plays an significant role in the maintenance of structural and functional integrity of the ER and mitochondria as well as pancreatic b-cells in type 2 diabetes (Bernales et al., 2007). However, autophagy is an important type of programmed cell death called type 2 programmed cell death or autophagic cell death under certain circumstances. Autophagic cell death is characterized by the accumulation of autophagic vesicles, which distinguishes it from type 1 programmed cell death or apoptosis (Tsujimoto and Shimizu, 2005). Nevertheless, whether autophagy plays a protective or harmful role in cell survival and death, it is undoubtedly essential for the pathophysiology of type 2 diabetes. Palmitate (PA) is used to enhance autophagy and elicit apoptosis in INS-1 b-cells. Recent in vitro studies showed a 10% increase in apoptosis when INS-1 cells were treated with 3-methyadenine (3-MA), an autophagy inhibitor and a 26% increase in apoptosis Q2 when cells were treated with 0.5 mmol l 1 PA + 3-MA for 24 h. In addition, 3-MA significantly reduced INS-1 cell viability by 54% and 40% by treating with 3-MA alone and 3-MA plus PA, respectively (Jing Yin et al., 2013). In vivo, it has been reported that Atg7f/f:RIP-Cre mice (Atg7-deficient mice) had significantly higher nonfasting glucose levels and severely impaired glucose tolerance compared with Atg7f/f mice fed a high-fat diet for 12 weeks (Fujitani et al., 2009). These results indicate that autophagy, a protein degradation system, is indispensable in the maintenance of normal cell function and survival (Fujitani et al., 2009). However, Fujimoto et al. indicted that activation of the autophagy pathway can increase b-cell death that occurs with reduced expression of pancreas duodenal homeobox 1 (Pdx1), and inhibition of autophagy prolonged cell survival and delayed cell death. They demonstrated that increased autophagy was present in Pdx1-reduced MIN6 cells and in Pdx1+/ b cells in vivo. The autophagy inhibitor 3-MA delays Pdx1 KD-induced MIN6 cell death, but 3-MA-treated Pdx1 KD MIN6 cells ultimately die by caspase3-dependent cell death. Thus, an increase in autophagy appears to occur early, and this is followed by an increase in apoptosis (Fujimoto et al., 2009). In vivo, Pdx1+/ mice were crossed to Becn1+/ mice to study the functional significance of increased autophagy in Pdx1+/ b cells, and autophagy was reduced in this model compared with wild-type mice. After 1 week on the high fat diet, the blood glucose concentrations of Pdx1+/ Becn1+/ mice were significantly decreased after the intraperitoneal administration of glucose when compared with blood glucose levels in Pdx1+/ mice (Fujimoto et al., 2009). Moreover, Becn1 is a key mediator of autophagosome formation and is the mammalian orthologue of the yeast Atg6/Vps30 gene and a regulator of the class III phosphoinositide 3-kinase complex involved in autophagosome formation


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b cells respond to insulin resistance by increasing b-cell mass and insulin secretion (hyperinsulinemia). When islets are unable to sustain b-cell compensation during insulin resistance, a relative insulin deficiency develops in addition to insulin resistance, and overt type 2 diabetes occurs (Prentki and Nolan, 2006). Serum long-chain FFA released from visceral fat of obese subjects constitutes one of the major culprits in the pathogenesis of insulin resistance and is a prerequisite for the development of type 2 diabetes. In vitro studies indicate that exposing b-cells to FFAs, such as palmitate and stearic acids, induces autophagy. Furthermore, inhibition of autophagosome formation augments FFA-induced b-cell death (Choi et al., 2009). Based on these observations, we can conclude that induced b-cell autophagy is an adaptive response against increased insulin resistance in which autophagy acts as a protective mechanism. In contrast, Zhou et al. showed that inhibition of autophagy prevented insulin receptor down-regulation induced by ER stress and that this insulin receptor down-regulation may play a critical role in obesity-induced insulin resistance (Zhou et al., 2009). The underlying mechanisms of ER stress-induced inhibition of insulin signaling remain elusive; however, some evidence suggests that serine phosphorylation of IRS-1/2 by JNK is critical in ER stressinduced insulin resistance (Ozcan et al., 2004, 2006). When 3T3-L1 adipocytes were treated with 3-MA, an autophagy inhibitor, the induction of autophagy by thapsigargin (a chemical known to induce ER stress by inhibiting ER Ca2+ATPase) was suppressed, and insulin receptor was protected from ER stress-induced degradation. However, this study also found that 3-MA was unable to restore insulin-stimulated Akt phosphorylation in the thapsigargin – treated cells. These results indicated that although blocking autophagy prevents ER stress-induced insulin receptor down-regulation, the majority of the ‘‘rescued’’ insulin receptor may already be defective, most likely as a result of ER stress-induced misfolding (Zhou et al., 2009). Autophagy may play different roles in different cells, but undoubtedly, it is closely associated with insulin resistance and may regulate essential processes of type 2 diabetes insulin resistance.

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5. Autophagy and ER stress

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To study the role of autophagy in diabetes, we studied the ER because ER distention was observed in autophagy-deficient b-cells, and ER stress is important in the pathogenesis of diabetes. Analysis of the ultrastructural changes using electron microscopy showed cisternal distension of rough ER even in apparently normal-looking b cells of Atg7Db-cell mice at lower magnifications. This morphological finding implied the existence of ER stress, which could lead to b-cell death and malfunction (Quan et al., 2012). Based on these results, we further analyzed the structure of autophagosomes. Initially, recent studies suggested the ER as the main source of autophagosomal membrane and the starting structure of autophagosomes. Utilizing 3D electron tomography and immunolabeling on thin-slice sections using antibodies against ER (anti protein disulfide isomerase) and isolation membrane (anti-Atg16L) showed that protein disulfide isomerase, an ER-resident protein, exists both outside and inside the isolation membrane structure. This suggests that the isolation membrane is formed when the autophagosome cradle is stacked between two ER membranes, and this structure is called the omegasome (Hamasaki and Yoshimori, 2010). Aggregated or terminally misfolded proteins in the ER lumen may be cleared by an autophagy, and the omegasomes double FYVE-containing protein 1 could be considered molecular markers of this process (Hamasaki and Yoshimori, 2010; Smith et al., 2011). Usually excess amounts of unfolded or misfolded proteins are cleared through ubiquitination and

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proteasomal degradation in the cytoplasm in a process called ERAD. If the amount of unfolded and misfolded proteins exceeds the capacity of the ERAD system, the proteins start to aggregate in the ER and trigger ER-stress-mediated cell death with caspase12 activation, which is specifically localized within the ER (Smith et al., 2011). Some studies suggest that PolyQ functions as a cytoplasmic aggregate of misfolded proteins, stimulates ER stress signals and induces ER-stress-mediated cell death by caspase12 activation in mouse cells. This process also induces the upregulation of Atg5, Atg7, Atg12, and Atg16 mRNAs via eIF2a phosphorylation, and this increased expression was inhibited by the eIF2aA/A mutation, which replaced a Ser able to be phosphorylated with Ala. Kouroku et al. also examined PolyQ72-induced LC3 conversion, CHOP, and Atg12 upregulation, and found these molecules to be inhibited by dominant-negative-PERK and the eIF2aA/A mutation (Kouroku et al., 2007). The results suggested that Atg5–Atg12–Atg16 complex-dependent LC3 conversion that leads to autophagy also inhibits polyQ72-induced ER-stress mediated cell death by caspase12 activation and degradation of cytoplasmic polyQ72 aggregates (Kouroku et al., 2007). In conclusion, these findings indicated that autophagy formation is a cellular defense mechanism against polyQ72-induced ER-stress-mediated cell death, with PERK/eIF2a phosphorylation is involved in polyQ72-induced LC3 conversion. However, these results have not been studied in pancreatic b cells, and it is still unclear whether the same processes could occur in this population and whether this would affect glucose homeostasis. These results may assist in the development of new therapies for type 2 diabetes. Recently, some studies showed that DAPK is a necessary regulator of autophagy and is activated by signals such as ER stress and cytokines. Several studies have mapped DAPK function to distinct stages in autophagy signaling, including interactions with the Beclin 1-phosphatidylinositol 3-kinase [PtdIns(3)K] complex, which is crucial in the early stages of autophagosome formation, and with the LC3-binding protein, MAP1B, which may regulate vesicle trafficking (Bialik and Kimchi, 2010). However, inflammatory cytokines may be essential for the mechanisms of ER stress and autophagy in type 2 diabetes, which has not been very clear until now.

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6. The connections among ER stress, inflammation and autophagy in type 2 diabetes

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Inflammaging refers to a low-grade pro-inflammatory phenotype that accompanies aging in mammals. The aging process is associated with a decline in autophagic capacity that contributes to cellular survival against nutrient deprivation and the turnover of damaged organelles. This phenomenon is related to type 2 diabetes, tumorigenesis, neurodegenerative diseases, cardiomyopathy and many other autoimmune and inflammatory diseases (Levine and Kroemer, 2008; Mizushima et al., 2008; Eskelinen and Saftig, 2009). First, there is some experimental evidence that suggests that MCP1 triggers the expression of a recently identified novel zinc-finger protein that induces ER stress that leads to autophagy. This protein also controls the inflammatory response by negatively regulating NF-kB, a master regulator of inflammation, via deubiquitination, and these effects may be associated with type 2 diabetes (Kolattukudy and Niu, 2012). Secondly, NLRP3 inflammasomes are molecular platforms activated by infection or stress (e.g., ER stress, ROS) that regulate the activity of caspase1, which cleaves the inactive precursors of IL1b and IL18 to stimulate their secretion (De Nardo and Latz, 2011). Experiments in macrophages have shown that the induction of NLRP3 inflammasomes triggers activation of the G protein RalB and autophagosome formation (Shi et al., 2012). Stimulation of inflammasomes led to autophagosome for-

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Cell stress (high glucose; cytokines; FFA ) Βcell death

IL1β

βcell

ER stress LC3

IRE1α

TXNIP

ChREBP ATF5

Fig. 1. TXNIP is induced by ER stress activates IL-1b production by the NLRP3 inflammasome, and mediates ER stress-mediated b-cell death. The inflammatory response is frequently triggered as a consequence of ER stress. A chronic excess of metabolic factors (such as lipids, glucose and cytokines) can cause ER stress. TXNIP upregulation by ER stress is mediated by the IRE1a and PERK-eIF2a in b-cells. Additionally, TXNIP is regulated by ChREBP and ATF5. PERK mediated eIF2a phosphorylation directs ATF5 protein translation and ChREBP expression.

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mation, which was initially independent of IL1b, ASC or caspase1 production and may occur during type 2 diabetes (Levine et al., 2011; Wen et al., 2011). Autophagy was able to capture and degrade inflammasomes via inflammasome ubiquitination, which led to the recruitment of p62 and LC3 (Levine et al., 2011). However, the activation of autophagy by inflammatory signals in b-cells remains poorly understood and must be further explored.

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7. Conclusion

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The prevalence of type 2 diabetes, an inflammatory disease, is growing rapidly worldwide. Prolonged or chronic inflammation can exacerbate tissue damage and is implicated in the development of type 2 diabetes (Dinarello, 2011). The inflammatory response is frequently triggered as a consequence of ER stress caused by metabolic problems or by the accumulation of misfolded proteins (Hotamisligil, 2010). From this review, we learned that ER stress, through the PERK and IRE1 pathways, activates IL1b production by the NLRP3 inflammasome (Fig. 1). LC3 conversion to LC3-II is also mediated by PERK/eIF2a phosphorylation, which suggests that PERK-mediated ER stress is essential for both the formation of inflammatory cytokines and the process of autophagy (Gonzalez et al., 2011; Oslowski et al., 2012). A current study indicated that Atg7-induced Cathepsin B overexpression resulted in an unexpected significant increase in proinflammatory chemokine and IL1b production in INS-1 (823/13) cells, which ultimately revealed the relationship between autophagy and inflammation in type 2 diabetes (Li et al., 2013). In light of the recent findings that connect autophagy and inflammasome regulation, it remains to be determined whether autophagy plays a protective or harmful role in diabetes. Although major components of the metabolic inflammasome promote autophagy, the induction of autophagy by this signaling complex would be expected to serve as a negative-feedback mechanism that limits ER stress and disease progression. Based on the above consideration, we support the establishment of a suitable method for monitoring the level of intracellular autophagy and inflammatory cytokines in prediabetic patients. This process could be helped by the elucidation of the molecular mechanisms of autophagy and inflammation, and the proper manipulation of these pathways could help to establish a novel therapeutic strategy for type 2 diabetes.

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Declaration of Interest

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Funding

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This work was supported by the National Natural Science Foundation of China (81370929), the Novo Nordisk China Diabetes Young Scientific Talent Research Funding (2013) program, a Grant from the Health Department of Heilongjiang Province (2012-540) and a Grant from the Chinese Medical Association of Clinical Medicine special funds for scientific research projects.

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Uncited reference

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No potential conflicts of interest were disclosed.

Mandrup-Poulsen et al. (2010).

Q3

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Acknowledgments

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This work was supported by the National Natural Science Foun- Q4 dation of China (81370929), the Novo Nordisk China Diabetes Q5 Young Scientific Talent Research Funding (2013) program, a Grant from the Health Department of Heilongjiang Province (2012-540) and a Grant from the Chinese Medical Association of Clinical Medicine special funds for scientific research projects.

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Appendix A. Supplementary data

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Supplementary data associated with this article can be found, in the online version, at http://dx.doi.org/10.1016/j.ygcen.2014. 09.006.

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autophagy pathway is involved in cholesterol-induced pancreatic β-cell injury.

The deadly connection between endoplasmic reticulum, Ca2+, protein synthesis, and the endoplasmic reticulum stress response in malignant glioma cells.

Endoplasmic reticulum stress is the crossroads of autophagy, inflammation, and apoptosis signaling pathways and participates in liver fibrosis.

Palmitate induces endoplasmic reticulum stress and autophagy in mature adipocytes: implications for apoptosis and inflammation.

Endoplasmic reticulum stress is involved in restraint stress-induced hippocampal apoptosis and cognitive impairments in rats.

Role of Endoplasmic Reticulum Stress, Autophagy, and Inflammation in Cardiovascular Disease.

Interplay of endoplasmic reticulum stress and autophagy in neurodegenerative disorders.

Lipotoxic endoplasmic reticulum stress, β cell failure, and type 2 diabetes mellitus.

Bufalin induces the interplay between apoptosis and autophagy in glioma cells through endoplasmic reticulum stress.

A molecular web: endoplasmic reticulum stress, inflammation, and oxidative stress.

Is glycemic control modulating endoplasmic reticulum stress in leukocytes of type 2 diabetic patients?

Type 2 diabetes mellitus induces congenital heart defects in murine embryos by increasing oxidative stress, endoplasmic reticulum stress, and apoptosis.

Crosstalk Between Endoplasmic Reticulum Stress, Oxidative Stress, and Autophagy: Potential Therapeutic Targets for Acute CNS Injuries.

Alzheimer's disease and type 2 diabetes: what is the connection?

Endoplasmic reticulum stress is involved in the lidocaine-induced apoptosis in SH-SY5Y neuroblastoma cells.

Endoplasmic Reticulum Stress Interacts With Inflammation in Human Diseases.

The endoplasmic reticulum-sarcoplasmic reticulum connection: distribution of endoplasmic reticulum markers in the sarcoplasmic reticulum of skeletal muscle fibers.

Endoplasmic Reticulum Stress and Microvascular Endothelial Dysfunction in Diabetes.

Endoplasmic reticulum stress eIF2α-ATF4 pathway-mediated cyclooxygenase-2 induction regulates cadmium-induced autophagy in kidney.

Endoplasmic reticulum and autophagy in rat hepatocytes.

Endoplasmic reticulum stress-induced PCD and caspase-like activities involved.

Endoplasmic reticulum stress-mediated apoptotic pathway is involved in corpus luteum regression in rats.

Effect of Liraglutide on endoplasmic reticulum stress in diabetes.

Genetic association and functional role of Crohn disease risk alleles involved in microbial sensing, autophagy, and endoplasmic reticulum (ER) stress.