脂肪细胞的内质网应激

肥胖常伴有炎性反应、胰岛素抵抗和全身代谢紊乱,脂肪细胞内质网应激在其中发挥莺要作用,内质网有合成、折叠和运输蛋白质的作用.脂肪细胞内质网还可以合成脂肪和感受营养物质的变化.在肥胖状态下,脂肪细胞受到营养过剩、细胞内胰岛素抵抗及氧化应激等因素的刺激后,内质网的生理功能发生紊乱.导致内质网应激,内质网应激的表现主要是未折叠蛋白质反应被激活以后,可以通过炎性反应和氧化应激等通路加重胰岛素抵抗.导致全身代谢紊乱.维持和增强内质网功能可能是改善胰岛素抵抗有效的方法之一,一些可以缓解内质网应激的化学分子伴侣是有前景的治疗药物.     

Nitric oxide-induced apoptosis in pancreatic beta cells is mediated by the endoplasmic reticulum stress pathway

Excessive nitric oxide (NO) production in cytokine-activated beta cells has been implicated in beta cell disruption in type 1 diabetes. beta cells are very vulnerable to NO-induced apoptosis. However, the mechanism underlying this phenomenon is unclear. Low concentrations of NO that lead to apoptosis apparently do not cause severe DNA damage in mouse MIN6 beta cells. CHOP, a C/EBP homologous protein that is induced by endoplasmic reticulum (ER) stress and plays a role in growth arrest and cell death, was induced by a NO donor, S-nitroso-N-acetyl-D,L-penicillamine (SNAP). SNAP increased cytosolic Ca(2+), and only agents depleting ER Ca(2+) induced CHOP expression and led to apoptosis, suggesting that NO depletes ER Ca(2+). Overexpression of calreticulin increased the Ca(2+) content of ER and afforded protection to cells against NO-mediated apoptosis. Furthermore, pancreatic islets from CHOP knockout mice showed resistance to NO. We conclude that NO depletes ER Ca(2+), causes ER stress, and leads to apoptosis. Thus, ER Ca(2+) stores are a new target of NO, and the ER stress pathway is a major mechanism of NO-mediated beta cell apoptosis.     

Trafficking-defective mutant PROKR2 cycles between endoplasmic reticulum and Golgi to attenuate endoplasmic reticulum stress

G protein–coupled receptors (GPCRs) play crucial roles in numerous physiological and pathological processes. Mutations in GPCRs that result in loss of function or alterations in signaling can lead to inherited or acquired diseases. Herein, studying prokineticin receptor 2 (PROKR2), we initially identify distinct interactomes for wild-type (WT) versus a mutant (P290S) PROKR2 that causes hypogonadotropic hypogonadism. We then find that both the WT and mutant PROKR2 are targeted for endoplasmic reticulum (ER)-associated degradation, but the mutant is degraded to a greater extent. Further analysis revealed that both forms can also leave the ER to reach the Golgi. However, whereas most of the WT is further transported to the cell surface, most of the mutant is retrieved to the ER. Thus, the post-ER itinerary plays an important role in distinguishing the ultimate fate of the WT versus the mutant. We have further discovered that this post-ER itinerary reduces ER stress induced by the mutant PROKR2. Moreover, we extend the core findings to another model GPCR. Our findings advance the understanding of disease pathogenesis induced by a mutation at a key residue that is conserved across many GPCRs and thus contributes to a fundamental understanding of the diverse mechanisms used by cellular quality control to accommodate misfolded proteins.

Gprotein–coupled receptors (GPCRs), also known as seven transmembrane domain receptors, are the largest protein family encoded by the human genome. They serve many cellular functions and are the largest class of therapeutic drug targets (1–3). Mutations in GPCRs are being identified in many disease settings, including patients with neuroendocrine diseases (4). The understanding of abnormal phenotypes caused by these mutations has contributed to our knowledge of the pathophysiology of human diseases.Several GPCRs, including gonadotropin-releasing hormone receptor (GnRHR), kisspeptin receptor, and tachykinin receptor 3, have roles in the reproductive neuroendocrine system. Mutations in these GPCRs lead to normosmic hypogonadotropic hypogonadism (HH) and Kallmann syndrome (KS), which are characterized by GnRH deficiency (5, 6). Prokineticin receptor 2 (PROKR2) and its ligand PROK2 have functional roles in multiple biological processes, including intestinal contraction, circadian rhythms, vascular and reproductive function, and the development of the olfactory system (7–13). Prokr2-deficient mice exhibit hypoplasia of the olfactory bulbs and abnormal migration of GnRH neurons as well as immature reproductive organs in both sexes (10). To date, over 20 missense mutations in the PROKR2 gene have been reported in patients with HH or KS (6, 14).Human disorders associated with GPCR mutations—for example, retinitis pigmentosa, ovarian dysgenesis, and nephrogenic diabetes as well as HH and KS—can occur as a result of defects in GPCR folding and trafficking (15). Over 50% of PROKR2 mutations identified in patients with HH and KS (11 of 21 mutations tested by functional assays) show impaired cell surface expression (6, 8, 16–19). Pharmacological chaperones have been used to rescue cell surface transport and function of some misfolded proteins, including GPCRs (20–22). For example, A457, an antagonist of PROKR2, has been used to restore cell surface expression and function of the P290S PROKR2 mutant, while treatment with 10% glycerol significantly increased cell surface expression and the signaling of P290S and W178S PROKR2 mutants (23). Thus, because cell surface expression is so critical for the function of many GPCRs, a better understanding of the different ways that mutations can alter the surface expression of GPCRs has been an important ongoing goal.In the endoplasmic reticulum (ER), a quality control system ensures that only correctly folded proteins are transported out of this compartment, while misfolded proteins are targeted for degradation. Protein-folding chaperones in the ER such as BiP (also known as GRP78 or HSPA5) prevent protein aggregation by promoting the refolding of misfolded proteins. Exemplifying this role, studies have shown that misfolded mutant GPCRs have increased association with ER chaperones compared to wild-type (WT) GPCRs (24–26). When refolding is unsuccessful, protein aggregation occurs in the ER, which evokes ER stress, leading to the activation of the unfolded protein response (UPR). The UPR system aims to restore normal cellular function and reduce ER stress. Failure to do so leads to major diseases, including neurodegenerative disorders, metabolic disorders, and cancer (27–29). Therefore, the ability to maintain a manageable level of ER stress is essential for human health.Misfolded proteins can be targeted for degradation following the activation of the UPR system in multiple ways. A major way occurs through ER-associated degradation (ERAD), which involves the retro-translocation of misfolded proteins out of the ER followed by their ubiquitination for targeting to the proteasome. Another way involves transport out of the ER followed by targeting to the lysosome for degradation. It has also been demonstrated that mutations in GnRHR, identified in patients with HH, result in degradation that combines elements of both proteasomal and autophagic degradation (30).With respect to the ERAD-based mechanism, it has been observed that some proteins are transported out of the ER and then retrieved from the Golgi back to the ER for degradation (31, 32). Why this occurs has been unclear. Here, we have studied in detail how a mutation at a residue in PROKR2, which is conserved across many GPCRs, affects its intracellular transport to alter function. Our results advance the understanding of disease pathogenesis induced by a key GPCR mutation and also contribute to a fundamental understanding of the different ways that misfolded proteins can be managed by the cell to maintain homeostasis.     

糖尿病性心肌病与内质网应激的关系

内质网应激（ERS）在维持内质网稳态的适应性应答中发挥着关键作用。近年来关于糖尿病(DM)的研究越来越多,研究显示在糖尿病性心肌病(DCM)早期,高血糖症、过多的游离脂肪酸以及炎症等多种致病因素均可导致ERS,ERS在DM的发展及其并发症形成中发挥着极为重要的作用。本文对ERS的经典通路、内质网功能紊乱与细胞自噬、凋亡的联系以及由于内质网功能紊乱导致机体产生胰岛素抵抗和DCM的过程进行了综述。     

巨噬细胞内质网应激与糖尿病大血管病变

糖尿病大血管病变是糖尿病慢性并发症之一,其主要病理特征是动脉粥样硬化.近年来的研究表明,巨噬细胞内质网应激与大血管动脉粥样硬化的发生、发展密切相关,糖尿病状态下的胰岛素抵抗,糖、脂代谢异常以及慢性炎性反应可诱导巨噬细胞内质网应激,从而促进动脉粥样硬化,最终加重了糖尿病大血管病变.关注巨噬细胞内质网应激在糖尿病大血管病变中的作用,将为抗动脉粥样硬化、治疗糖尿病大血管病变提供一个新方向.     

内质网应激与慢性阻塞性肺疾病

内质网应激(endoplasmic reticulum stress,ERS)反应是由于多种应急原引起的细胞内质网功能障碍,导致内质网腔内错误折叠、未折叠蛋白聚集和钙离子平衡紊乱,细胞为恢复内质网功能而进行的一系列调节反应.ERS广泛参与多种疾病的发病机制,是一把介导细胞适应性生存和凋亡/自噬的双刃剑.肺结构细胞凋亡是慢性阻塞性肺疾病发生、发展的重要机制.近年的研究表明,ERS在慢性阻塞性肺疾病肺结构细胞凋亡机制中发挥了重要的作用.     

Alcohol abuse, endoplasmic reticulum stress and pancreatitis

Alcohol abuse is a common cause of both acute and chronic pancreatitis. There is a wide spectrum of pancreatic manifestations in heavy drinkers from no apparent disease in most individuals to acute inflammatory and necrotizing pancreatitis in a minority of individuals with some progressing to chronic pancreatitis characterized by replacement of the gland by fibrosis and chronic inflammation. Both smoking and African-American ethnicity are associated with increased risk of alcoholic pancreatitis. In this review we describe how our recent studies demonstrate that ethanol feeding in rodents causes oxidative stress in the endoplasmic reticulum (ER) of the digestive enzyme synthesizing acinar cell of the exocrine pancreas. This ER stress is attenuated by a robust unfolded protein response (UPR) involving X-box binding protein-1 (XBP1) in the acinar cell. When the UPR activation is prevented by genetic reduction in XBP1, ethanol feeding causes significant pathological responses in the pancreas. These results suggest that the reason most individuals who drink alcohol heavily do not get significant pancreatic disease is because the pancreas mounts an adaptive UPR to attenuate the ER stress that ethanol causes. We hypothesize that disease in the pancreas results when the UPR is insufficiently robust to alleviate the ER stress caused by alcohol abuse.     

Neuronal nitric oxide synthase protects the pancreatic beta cell from glucolipotoxicity-induced endoplasmic reticulum stress and apoptosis

Aims/hypothesis  

Cytokines stimulate nitric oxide production in pancreatic beta cells, leading to endoplasmic reticulum (ER) stress and apoptosis. Treatment of beta cells with glucose and NEFA induces nitric oxide synthase (NOS) as well as ER stress. However, the role of NO in glucolipotoxicity-induced ER stress in beta cells is not clear.     

内质网应激与心血管疾病的研究进展

内质网(ER)是真核细胞最主要的膜性结构,是细胞内重要生理过程发生的关键细胞器。在多种内外因素的作用下,ER的稳态受到破坏,导致蛋白质加工运输受阻,未折叠蛋白或错误折叠蛋白在ER腔内聚集,形成内质网应激(ERS),并触发未折叠蛋白反应(UPR)。适度的ERS通过UPR信号通路减少蛋白质合成、促进蛋白质降解、增加协助蛋白质折叠的分子伴侣,最终缓解ER压力。但是,如果ERS过强或持续时间过长,超过细胞的自身调节能力时,UPR可启动细胞凋亡,亦可导致疾病。大量研究表明,ERS与多种心血管疾病(CVD)的发生发展密切相关。该综述主要阐述UPR在几种常见CVD中的研究进展和靶向UPR作为CVD的潜在治疗方法。     

Monogenic and syndromic diabetes due to endoplasmic reticulum stress

The endoplasmic reticulum (ER) lies at the crossroads of protein folding, calcium storage, lipid metabolism, and the regulation of autophagy and apoptosis. Accordingly, dysregulation of ER homeostasis leads to β-cell dysfunction in type 1 and type 2 diabetes that ultimately culminates in cell death. The ER is therefore an emerging target for understanding the mechanisms of diabetes mellitus that captures the complex etiologies of this multifactorial class of metabolic disorders. Our strategy for developing ER-targeted diagnostics and therapeutics is to focus on monogenic forms of diabetes related to ER dysregulation in an effort to understand the exact contribution of ER stress to β-cell death. In this manner, we can develop personalized genetic medicine for ERstress-related diabetic disorders, such as Wolfram syndrome. In this article, we describe the phenotypes and molecular pathogenesis of ERstress-related monogenic forms of diabetes.     

阿托伐他汀减轻巨噬细胞内质网应激及其介导的细胞凋亡

目的 探讨阿托伐他汀对7-酮胆固醇（7-KC）诱导的巨噬细胞内质网应激及细胞凋亡的影响。方法 AopE-/-小鼠左肾动脉和左颈总动脉联合部分结扎建立颈动脉易损斑块模型。采用HE染色方法观察斑块病理学改变,用免疫荧光结合激光扫描共聚焦显微镜技术检测斑块中内质网应激(ER stress)相关蛋白CHOP及磷酸化PERK（p-PERK）的表达。体外培养小鼠巨噬细胞RAW264.7,给予7-KC、H2O2或联合阿托伐他汀处理后,蛋白质免疫印迹方法（Western blot）测定ER stress相关蛋白CHOP、p-PERK 、XBP-1s及凋亡相关蛋白cleaved caspase-3的表达。结果 AopE-/-小鼠颈动脉易损斑块局部ER stress相关蛋白CHOP的表达及PERK磷酸化水平明显上调;7-KC可诱导小鼠巨噬细胞ER stress,进而诱导细胞凋亡;同时,氧化应激诱导剂H2O2也可通过诱导小鼠巨噬细胞ER stress介导细胞凋亡;而阿托伐他汀可抑制7-KC和H2O2诱导的巨噬细胞ER stress及其介导的细胞凋亡。结论 ER stress可能参与AS易损斑块的形成;阿托伐他汀可通过减少细胞内氧化应激的水平,减轻巨噬细胞ER stress,从而抑制细胞凋亡。     

Antioxidants reduce endoplasmic reticulum stress and improve protein secretion

Protein misfolding in the endoplasmic reticulum (ER) contributes to the pathogenesis of many diseases. Although oxidative stress can disrupt protein folding, how protein misfolding and oxidative stress impact each other has not been explored. We have analyzed expression of coagulation factor VIII (FVIII), the protein deficient in hemophilia A, to elucidate the relationship between protein misfolding and oxidative stress. Newly synthesized FVIII misfolds in the ER lumen, activates the unfolded protein response (UPR), causes oxidative stress, and induces apoptosis in vitro and in vivo in mice. Strikingly, antioxidant treatment reduces UPR activation, oxidative stress, and apoptosis, and increases FVIII secretion in vitro and in vivo. The findings indicate that reactive oxygen species are a signal generated by misfolded protein in the ER that cause UPR activation and cell death. Genetic or chemical intervention to reduce reactive oxygen species improves protein folding and cell survival and may provide an avenue to treat and/or prevent diseases of protein misfolding.     

Biology of endoplasmic reticulum stress in the heart

The endoplasmic reticulum (ER) is a multifunctional intracellular organelle supporting many processes required by virtually every mammalian cell, including cardiomyocytes. It performs diverse functions, including protein synthesis, translocation across the membrane, integration into the membrane, folding, posttranslational modification including N-linked glycosylation, and synthesis of phospholipids and steroids on the cytoplasmic side of the ER membrane, and regulation of Ca(2+) homeostasis. Perturbation of ER-associated functions results in ER stress via the activation of complex cytoplasmic and nuclear signaling pathways, collectively termed the unfolded protein response (UPR) (also known as misfolded protein response), leading to upregulation of expression of ER resident chaperones, inhibition of protein synthesis and activation of protein degradation. The UPR has been associated with numerous human pathologies, and it may play an important role in the pathophysiology of the heart. ER stress responses, ER Ca(2+) buffering, and protein and lipid turnover impact many cardiac functions, including energy metabolism, cardiogenesis, ischemic/reperfusion, cardiomyopathies, and heart failure. ER proteins and ER stress-associated pathways may play a role in the development of novel UPR-targeted therapies for cardiovascular diseases.     

Hyperhomocysteinemia, endoplasmic reticulum stress, and alcoholic liver injury

Deficiencies in vitamins or other factors (B6, B12, folic acid, betaine) and genetic disorders for the metabolism of the non-protein amino acid-homocysteine (Hcy) lead to hyperhomocysteinemia (HHcy). HHcy is an integral component of several disorders including cardiovascular disease, neurodegeneration, diabetes and alcoholic liver disease. HHcy unleashes mediators of inflammation such as NFkB, IL-1β, IL-6, and IL-8, increases production of intracellular superoxide anion causing oxidative stress and reducing intracellular level of nitric oxide (NO), and induces endoplasmic reticulum (ER) stress which can explain many processes of Hcy-promoted cell injury such as apoptosis, fat accumulation, and inflammation. Animal models have played an important role in determining the biological effects of HHcy. ER stress may also be involved in other liver diseases such as a1-antitrypsin (a1-AT) deficiency and hepatitis C and/or B virus infection. Future research should evaluate the possible potentiative effects of alcohol and hepatic virus infection on ER stress-induced liver injury, study potentially beneficial effects of lowering Hcy and preventing ER stress in alcoholic humans, and examine polymorphism of Hcy metabolizing enzymes as potential risk-factors for the development of HHcy and liver disease.     

Metallothionein alleviates oxidative stress-induced endoplasmic reticulum stress and myocardial dysfunction

Oxidative stress and endoplasmic reticulum (ER) stress have been implicated in cardiovascular diseases although the interplay between the two is not clear. This study was designed to examine the influence of oxidative stress through glutathione depletion on myocardial ER stress and contractile function in the absence or presence of the heavy metal scavenger antioxidant metallothionein (MT). FVB and MT overexpression transgenic mice received the GSH synthase inhibitor buthionine sulfoximine (BSO, 30 mM) in drinking water for 2 weeks. Oxidative stress, ER stress, apoptosis, cardiac function and ultrastructure were assessed using GSH/GSSG assay, reactive oxygen species (ROS), immunoblotting, caspase-3 activity, Langendorff perfused heart function (LVDP and ± dP/dt), and transmission electron microscopy. BSO led to a robust decrease in the GSH/GSSG ratio and increased ROS production, consolidating oxidative stress. Cardiac function and ultrastructure were compromised following BSO treatment, the effect of which was obliterated by MT. BSO promoted overt ER stress as evidenced by upregulated BiP, calregulin, phospho-IRE1α and phospho-eIF2α without affecting total IRE1α and eIF2α. BSO treatment led to apoptosis manifested as elevated expression of CHOP/GADD153, caspase-12 and Bax as well as caspase-3 activity, reduced Bcl-2 expression and JNK phosphorylation, all of which was ablated by MT. Moreover, both antioxidant N-acetylcysteine and the ER stress inhibitor tauroursodeoxycholic acid reversed the oxidative stress inducer menadione-elicited depression in cardiomyocyte contractile function. Taken together, these data suggested that ER stress occurs likely downstream of oxidative stress en route to cardiac dysfunction.     

内质网应激与胰岛素抵抗发生机制的关系

胰岛素抵抗(IR)与多种疾病密切相关,IR相关疾病的患病率逐年升高,近年来对于IR发生机制的研究成为热点.内质网应激(ERS)在IR的发生、发展中发挥重要作用,但其具体机制尚未完全明确.核因子kB(NF-kB)信号通路的激活与IR的发生、发展密切相关,并且与ERS间存在一定的联系,ERS可能通过激活NF-kB信号通路引起IR.