Diet-induced Endoplasmic Reticulum Stress Research Articles

Disodium Fumarate Alleviates Endoplasmic Reticulum Stress, Mitochondrial Damage, and Oxidative Stress Induced by the High-Concentrate Diet in the Mammary Gland Tissue of Hu Sheep

The long-term feeding of the high-concentrate diet (HC) reduced rumen pH and induced subacute rumen acidosis (SARA), leading to mammary gland tissue damage among ruminants. Disodium fumarate enhanced rumen bufferation and alleviated a decrease in rumen pH induced by the HC diet. Therefore, the purpose of this study was to investigate whether disodium fumarate could alleviate endoplasmic reticulum (ER) stress, mitochondrial damage, and oxidative stress induced by the high-concentrate diet in the mammary gland tissue of Hu sheep. In this study, 18 Hu sheep in mid-lactation were randomly divided into three groups: one fed with a low-concentrate diet (LC) diet, one fed with a HC diet, and one fed with a HC diet with disodium fumarate (AHC). Each sheep was given an additional 10 g of disodium fumarate/day. The experiment lasted for eight weeks. After the experiment, rumen fluid, blood, and mammary gland tissue were collected. The results show that, compared with the LC diet, the HC diet could reduce rumen pH, and the pH below 5.6 was more than 3 h, and the LPS content of blood and rumen fluid in HC the diet was significantly higher than in the LC diet. This indicates that the HC diet induced SARA in Hu sheep. However, the supplementation of disodium fumarate in the HC diet increased the rumen pH and decreased the content of LPS in blood and rumen fluid. Compared with the LC diet, the HC diet increased Ca2+ content in mammary gland tissue. However, the AHC diet decreased Ca2+ content. The HC diet induced ER stress in mammary gland tissue by increasing the mRNA and protein expressions of GRP78, CHOP, PERK, ATF6, and IRE1α. The HC diet also activated the IP3R-VDAC1-MCU channel and lead to mitochondrial damage by inhibiting mitochondrial fusion and promoting mitochondrial division, while disodium fumarate could alleviate these changes. In addition, disodium fumarate alleviated oxidative stress induced by the HC diet by activating Nrf2 signaling and reducing ROS production in mammary gland tissue. In conclusion, the supplementation of disodium fumarate at a daily dose of 10 g/sheep enhanced rumen bufferation by maintaining the ruminal pH above 6 and reduced LPS concentration in ruminal fluid and blood. This reaction avoided the negative effect observed by non-supplemented sheep that were fed with a high-concentrate diet involving endoplasmic reticulum stress, oxidative stress, and mitochondrial damage in the mammary gland tissue of Hu sheep.

Effect of Vitamin D Supplementation on High Fat Diet Induced Thyroid Dysfunction an RCT on Mouse Model

Background: Excess of dietary fats induce hyperlipidemia which causes endoplasmic reticulum (ER)stress. ER stress causes thyroid dysfunction and reduces serum thyroid hormone levels. Vitamin D supplementation increases thyroid hormone levels by reducing high fat diet induced ER stress, inflammation and autoimmunity. Objective: This study was planned to determine the effect of vitamin D supplementation on levels of serum free and total thyroxine in male albino mice. Methodology: In this randomized control trial, ninety healthy male albino mice were taken by consecutive, nonprobability sampling and randomly divided into three groups, each group containing 30 mice. Group A mice were given normal diet, Group B mice were given high fat diet, Group C mice were given high fat diet and vitamin D through oral gavage (100ng/kg/day) for 6 weeks. By the end of 6 weeks, blood sample of all mice was taken by cardiac puncture and serum was separated. Levels of serum total thyroxine and free thyroxine were measured by ELISA technique. Data was analyzed by using SPSS version 20. Results: In group B mice (high fat diet group) serum total and free thyroxine levels were reduced significantly (p=0.00) as compared to group A mice (normal diet group). In group C mice (high fat diet plus vitamin D group) serum total and free thyroxine levels were increased significantly (p=0.00) as compared to group B mice ( high fat diet group). The results of group C and A were also comparable and significant, with serum free and total thyroxine levels significantly high in group C (high fat diet+ vitamin D) as compared to group A(normal diet). Conclusion: Vitamin D supplemented mice on high fat diet had increased levels of serum total thyroxine, free thyroxine as compared to the group of mice on high fat diet only. Keywords: Vitamin D, High fat diet, Hyperlipidemia, Thyroxine.

Intercellular Transmission of Hepatic ER Stress in Obesity Disrupts Systemic Metabolism

Endoplasmic reticulum stress (ERS) has a pathophysiological role in obesity-associated insulin resistance. Yet, the coordinated tissue response to ERS remains unclear. Increased connexin 43 (Cx43)-mediated intercellular communication has been implicated in tissue-adaptive and -maladaptive response to various chronic stresses. Here, we demonstrate that in hepatocytes, ERS results in increased Cx43 expression and cell-cell coupling. Co-culture of ER-stressed "donor" cells resulted in intercellular transmission of ERS and dysfunction to ERS-naive "recipient" cells ("bystander response"), which could be prevented by genetic or pharmacologic suppression of Cx43. Hepatocytes from obese mice were able to transmit ERS to hepatocytes from lean mice, and mice lacking liver Cx43 were protected from diet-induced ERS, insulin resistance, and hepatosteatosis. Taken together, our results indicate that in obesity, the increased Cx43-mediated cell-cell coupling allows intercellular propagation of ERS. This novel maladaptive response to over-nutrition exacerbates the tissue ERS burden, promoting hepatosteatosis and impairing whole-body glucose metabolism.

An Endoplasmic Reticulum Stress-MicroRNA-26a Feedback Circuit in NAFLD.

Endoplasmic reticulum (ER) stress is an adaptive response to excessive ER demand and contributes to the development of numerous diseases, including nonalcoholic fatty liver disease (NAFLD), which is hallmarked by the accumulation of lipid within hepatocytes. However, the underlying mechanisms remain elusive. MicroRNAs (miRNAs) play an indispensable role in various stress responses, but their implications in ER stress have not yet been systemically investigated. In this study, we identify a negative feedback loop consisting of hepatic ER stress and miR-26a in NAFLD pathogenesis. Combining miRNA dot blot array and quantitative PCR, we find that miR-26a is specifically induced by ER stress in liver cells. This induction of miR-26a is critical for cells to cope with ER stress. In human hepatoma cells and murine primary hepatocytes, overexpression of miR-26a markedly alleviates chemical-induced ER stress, as well as palmitate-triggered ER stress and lipid accumulation. Conversely, deficiency of miR-26a exhibits opposite effects. Mechanistically, miR-26a directly targets the eukaryotic initiation factor 2α, a core ER stress effector controlling cellular translation. Intriguingly, miR-26a is reduced in the livers of patients with NAFLD. Hepatocyte-specific restoration of miR-26a in mice significantly mitigates high-fat diet-induced ER stress and hepatic steatosis. In contrast, deficiency of miR-26a in mice exacerbates high-fat diet-induced ER stress, lipid accumulation, inflammation and hepatic steatosis. Our findings suggest ER stress-induced miR-26a up-regulation as a regulator for hepatic ER stress resolution, and highlight the ER stress/miR-26a/eukaryotic initiation factor 2α cascade as a promising therapeutic strategy for NAFLD.

SAT-550 High Fat Diet-Induced ER Stress Affects Hypothyroidism

Recent studies revealed the emerging role of excess uptake of lipids in the development of hypothyroidism. Both dietary high-fat lard intake in rats and diet-induced obese mice displayed thyroid hypofunction (1,2). However, the underlying mechanism is largely unknown. We investigated the effect of high fat diet (HFD) on thyroid function and the role of endoplasmic reticulum (ER) in HFD induced hypothyroidism. Male Sprague Dawley rats were fed with HFD or control diet (CD) for 18 weeks. HFD rats showed increased serum TSH (n=15-16) and decreased T4 (n=15-16) compared to CD rats. Thyroid function was also evaluated by thyroid radioiodine uptake for 24 hours in vivo. Consistently, HFD decreased thyroid iodine uptake (-60%, n=5). To understand the molecular mechanism, we assessed the expression of proteins critical to thyroid production via Western blotting. Compared to CD, HFD rats showed significantly decreased thyroglobulin (Tg) (-40%, n=4), whereas TPO and NIS showed no difference. Meanwhile, luminal swelling of ER in thyroid epithelial cell of HFD rats was observed via transmission electron microscopy. Furthermore, We found PERK/eIF2a/ATF4 -- one of three traditional UPR pathways triggered under ER stress -- was activated in HFD rat thyroid glands compared to CD (n=4). In vitro, ER stress was induced in primary human thyrocytes by palmitate. Under the treatment of palmitate, Tg was degraded more rapidly than the control, indicating the activating of the ER-associated protein degradation pathway under ER stress. In addition, alleviation of ER stress by applying 4-phenyl butyric acid (4-PBA) in HFD rats or by the withdrawal of HFD improved the decrease of Tg (n=4-6) and thyroid function (n=3-6). In conclusion, we demonstrate that ER stress mediates the HFD induced hypothyroidism, probably by impairing the production of Tg, and attenuation of ER stress improves thyroid function. Reference: (1) Shao et al., Acta Pharmacol Sin 2014, 35: 1411-1420. (2) Lee et al., Endocrinology 2015, 156: 1181-1193. Conflict of interest: The authors declare they have no conflicts of interest. Sources of Research Support: National Natural Science Foundation of China (81430020, 81230018, 81600604, 81300644, and 81670720) and the Natural Science Foundation of Shandong Province (ZR2016HB07).

Hypothalamic kappa opioid receptor mediates both diet-induced and melanin concentrating hormone-induced liver damage through inflammation and endoplasmic reticulum stress.

The opioid system is widely known to modulate the brain reward system and thus affect the behavior of humans and other animals, including feeding. We hypothesized that the hypothalamic opioid system might also control energy metabolism in peripheral tissues. Mice lacking the kappa opioid receptor (κOR) and adenoviral vectors overexpressing or silencing κOR were stereotaxically delivered in the lateral hypothalamic area (LHA) of rats. Vagal denervation was performed to assess its effect on liver metabolism. Endoplasmic reticulum (ER) stress was inhibited by pharmacological (tauroursodeoxycholic acid) and genetic (overexpression of the chaperone glucose‐regulated protein 78 kDa) approaches. The peripheral effects on lipid metabolism were assessed by histological techniques and western blot. We show that in the LHA κOR directly controls hepatic lipid metabolism through the parasympathetic nervous system, independent of changes in food intake and body weight. κOR colocalizes with melanin concentrating hormone receptor 1 (MCH‐R1) in the LHA, and genetic disruption of κOR reduced melanin concentrating hormone–induced liver steatosis. The functional relevance of these findings was given by the fact that silencing of κOR in the LHA attenuated both methionine choline–deficient, diet‐induced and choline‐deficient, high‐fat diet–induced ER stress, inflammation, steatohepatitis, and fibrosis, whereas overexpression of κOR in this area promoted liver steatosis. Overexpression of glucose‐regulated protein 78 kDa in the liver abolished hypothalamic κOR‐induced steatosis by reducing hepatic ER stress. Conclusions: This study reveals a novel hypothalamic–parasympathetic circuit modulating hepatic function through inflammation and ER stress independent of changes in food intake or body weight; these findings might have implications for the clinical use of opioid receptor antagonists. (Hepatology 2016;64:1086‐1104)

Factors involved in the development of the metabolic syndrome. We are what we eat and what we are eating

A variety of metabolic and molecular changes in brain and adipose tissue play a critical role in the pathophysiology of life style-related metabolic diseases [1-3]. Even in obese subjects with insulin resistance in skeletal muscle and liver, insulin action on adipose tissue remains intact or rather exaggerated, resulting in considerable difficulties in weight reduction. Excess in circulating insulin thus causes body fat gain as well as ectopic lipid overload in liver, skeletal and cardiac muscle, pancreas and vasculature. The adipocyte hormone, leptin controls feeding behavior, augments fatty acid b-oxidation in the skeletal muscle and enhances whole body insulin sensitivity, thereby serving as a promising therapeutic candidate for the treatment of obesity-diabetes syndrome. However, the clinical application of leptin has been hampered by the notion that leptin does not fully exert its metabolic effects in subjects with fat diet-induced obesity [4-6]. It is important to note that the future risk of cardiometabolic diseases exists for infants with low birth weight/intrauterine growth retardation (IUGR). IUGR causes a premature leptin surge in the neonatal period in mice, and conceivably, through a number of epigenetic mechanisms, it may induce hypothalamic leptin resistance in adulthood. Mechanisms of non-genomic, intergenerational transmission of metabolic diseases from parents to children have also been unveiled in an expeditious fashion. The endoplasmic reticulum (ER) is an intracellular organelle involved in protein folding and apoptosis. The accumulation of misfolded proteins in the ER, termed as ER stress, is involved in the molecular pathophysiology of type 2 diabetes and metabolic syndrome. Our recent research in mice demonstrated that high fat diet-induced ER stress in the hypothalamus plays a pivotal role in the preference for fatty foods and resultant increase in body weight. We provided the first evidence that brown rice and its major component, g-oryzanol, ameliorate glucose dyshomeostasis in mice fed high-fat diet (HFD), accompanied by reduction of hypothalamic endoplasmic reticulum (ER) stress [7]. In my talk, I try to review the update of mechanisms of metabolic syndrome, with a particular focus on the molecular food sciences.

Soluble Epoxide Hydrolase Deficiency or Inhibition Attenuates Diet-induced Endoplasmic Reticulum Stress in Liver and Adipose Tissue

Soluble epoxide hydrolase (sEH) is a cytosolic enzyme whose inhibition has beneficial effects in cardiovascular, inflammatory, and metabolic diseases in murine models. Mice with targeted deletion or pharmacological inhibition of sEH exhibit improved insulin signaling in liver and adipose tissue. Herein, we assessed the role of sEH in regulating endoplasmic reticulum (ER) stress in liver and adipose tissue. We report that sEH expression was increased in the livers and adipose tissue of mice fed a high fat diet, the adipose tissue of overweight humans, and palmitate-treated cells. Importantly, sEH deficiency or inhibition in mice attenuated chronic high fat diet-induced ER stress in liver and adipose tissue. Similarly, pharmacological inhibition of sEH in HepG2 cells and 3T3-L1 adipocytes mitigated chemical-induced ER stress and activation of JNK, p38, and cell death. In addition, insulin signaling was enhanced in HepG2 cells treated with sEH substrates and attenuated in cells treated with sEH products. In summary, these findings demonstrate that sEH is a physiological modulator of ER stress and a potential target for mitigating complications associated with obesity.

Endoplasmic reticulum stress upregulates protein tyrosine phosphatase 1B and impairs glucose uptake in cultured myotubes.

Endoplasmic reticulum (ER) stress has been recognised as a common pathway in the development of obesity-associated insulin resistance. Protein tyrosine phosphatase 1B (PTP1B) is a negative regulator of insulin signalling and is localised on the ER membrane. The aim of the study was to investigate the cross-talk between ER stress and PTP1B. Leptin-deficient obese (ob/ob), Ptp1b (also known as Ptpn1) knockout and C57BL/6J mice were subjected to a high-fat or normal-chow diet for 20 weeks. ER stress was induced in cultured myotubes by treatment with tunicamycin. Immunohistochemistry and western blotting were used to assess proteins involved in the ER stress response. Myotube glucose uptake was determined by measuring 2-deoxy[(3)H]glucose incorporation. A high-fat diet induced ER stress and PTP1B expression in the muscle tissue of mice and these responses were attenuated by treatment with the ER chaperone tauroursodeoxycholic acid (TUDCA). Cultured myotubes exhibited increased levels of PTP1B in response to tunicamycin treatment. Silencing of Ptp1b with small interfering RNA (siRNA) or overexpression of Ptp1b with adenovirus construct failed to alter the levels of ER stress. Ptp1b knockout mice did not differ from the wild-type mice in the extent of tunicamycin-induced upregulation of glucose-regulated protein-78. However, tunicamycin-induced phosphorylation of eukaryotic initiation factor 2α and c-Jun NH(2)-terminal kinase-2 were significantly attenuated in the Ptp1b knockout mice. Treatment with TUDCA or silencing of PTP1B reversed tunicamycin-induced blunted myotube glucose uptake. Our data suggest that PTP1B is activated by ER stress and is required for full-range activation of ER stress pathways in mediating insulin resistance in the skeletal muscle.

Reducing endoplasmic reticulum stress does not improve steatohepatitis in mice fed a methionine- and choline-deficient diet

Endoplasmic reticulum (ER) stress has been implicated in the pathogenesis of nonalcoholic steatohepatitis. The ER stress response is activated in the livers of mice fed a methionine- and choline-deficient (MCD) diet, yet the role of ER stress in the pathogenesis of MCD diet-induced steatohepatitis is unknown. Using chemical chaperones on hepatic steatosis and markers of inflammation and fibrosis in mice fed a MCD diet, we aim to determine the effects of reducing ER stress. C57BL/6J mice were fed a MCD diet with or without the ER chemical chaperones 4-phenylbutyric acid (PBA) and tauroursodeoxycholic acid (TUDCA) for 2 wk. TUDCA and PBA effectively attenuated the ER stress response in MCD diet-fed mice, as evidenced by reduced protein levels of phosphorylated eukaryotic initiation factor 2α and phosphorylated JNK and suppression of mRNA levels of CCAAT/enhancer binding protein homologous protein, glucose-regulated protein 78 kDa, and X-box binding protein 1. However, PBA and TUDCA did not decrease MCD diet-induced hepatic steatosis. MCD diet-induced hepatic inflammation, as evidenced by increased plasma alanine aminotransferase and induction of hepatic TNFα expression, was also not reduced by PBA or TUDCA. PBA and TUDCA did not attenuate MCD diet-induced upregulation of the fibrosis-associated genes tissue inhibitor of metalloproteinase-1 and matrix metalloproteinase-9. ER chemical chaperones reduce MCD diet-induced ER stress, yet they do not improve MCD diet-induced hepatic steatosis, inflammation, or activation of genes associated with fibrosis. These data suggest that although the ER stress response is activated by the MCD diet, it does not have a primary role in the pathogenesis of MCD diet-induced steatohepatitis.

12/15-Lipoxygenase signaling in the endoplasmic reticulum stress response

Central obesity is associated with chronic inflammation, insulin resistance, β-cell dysfunction, and endoplasmic reticulum (ER) stress. The 12/15-lipoxygenase enzyme (12/15-LO) promotes inflammation and insulin resistance in adipose and peripheral tissues. Given that obesity is associated with ER stress and 12/15-LO is expressed in adipose tissue, we determined whether 12/15-LO could mediate ER stress signals. Addition of 12/15-LO lipid products 12(S)-HETE and 12(S)-HPETE to differentiated 3T3-L1 adipocytes induced expression and activation of ER stress markers, including BiP, XBP-1, p-PERK, and p-IRE1α. The ER stress inducer, tunicamycin, upregulated ER stress markers in adipocytes with concomitant 12/15-LO activation. Addition of a 12/15-LO inhibitor, CDC, to tunicamycin-treated adipocytes attenuated the ER stress response. Furthermore, 12/15-LO-deficient adipocytes exhibited significantly decreased tunicamycin-induced ER stress. 12/15-LO action involves upregulation of interleukin-12 (IL-12) expression. Tunicamycin significantly upregulated IL-12p40 expression in adipocytes, and IL-12 addition increased ER stress gene expression; conversely, LSF, an IL-12 signaling inhibitor, and an IL-12p40-neutralizing antibody attenuated tunicamycin-induced ER stress. Isolated adipocytes and liver from 12/15-LO-deficient mice fed a high-fat diet revealed a decrease in spliced XBP-1 expression compared with wild-type C57BL/6 mice on a high-fat diet. Furthermore, pancreatic islets from 12/15-LO-deficient mice showed reduced high-fat diet-induced ER stress genes compared with wild-type mice. These data suggest that 12/15-LO activity participates in ER stress in adipocytes, pancreatic islets, and liver. Therefore, reduction of 12/15-LO activity or expression could provide a new therapeutic target to reduce ER stress and downstream inflammation linked to obesity.