• Journal of Ginseng Research
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• v.40 no.4
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• pp.351-358
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• 2016

Background: This study was designed to investigate whether ginsenoside Rb1 (Rb1) and compound K (CK) ameliorated insulin resistance by suppressing endoplasmic reticulum (ER) stress-induced inflammation in adipose tissue. Methods: To induce ER stress, epididymal adipose tissue from mice or differentiated 3T3 adipocytes were exposed to high glucose. The effects of Rb1 and CK on reactive oxygen species production, ER stress, TXNIP/NLRP3 inflammasome activation, inflammation, insulin signaling activation, and glucose uptake were detected by western blot, emzyme-linked immunosorbent assay, or fluorometry. Results: Rb1 and CK suppressed ER stress by dephosphorylation of $IRE1{\alpha}$ and PERK, thereby reducing TXNIP-associated NLRP3 inflammasome activation in adipose tissue. As a result, Rb1 and CK inhibited IL-$1{\beta}$ maturation and downstream inflammatory factor IL-6 secretion. Inflammatory molecules induced insulin resistance by upregulating phosphorylation of insulin receptor substrate-1 at serine residues and impairing insulin PI3K/Akt signaling, leading to decreased glucose uptake by adipocytes. Rb1 and CK reversed these changes by inhibiting ER stress-induced inflammation and ameliorating insulin resistance, thereby improving the insulin IRS-1/PI3K/Akt-signaling pathway in adipose tissue. Conclusion: Rb1 and CK inhibited inflammation and improved insulin signaling in adipose tissue by suppressing ER stress-associated NLRP3 inflammation activation. These findings offered novel insight into the mechanism by which Rb1 and CK ameliorate insulin resistance in adipose tissue.

• Development and Reproduction
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• v.21 no.4
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• pp.407-415
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• 2017

In the present study, we investigated the role of binding immunoglobulin protein/glucose-regulated protein, 78-kDa (BIP/GRP78)-regulated endoplasmic reticulum (ER)-stress on meiotic maturation and cumulus cells expansion in porcine cumulus-oocyte complexes (COCs). Previously, it has been demonstrated that unfolded protein response (UPR)-related genes, such as molecules involved in ER-stress defense mechanisms, were expressed in matured oocytes and cumulus cells during in vitro maturation (IVM) of porcine oocytes. However, BIP/GRP78-mediated regulation of ER stress in porcine oocytes has not been reported. Firstly, we observed the effects of knockdown of BIP/GRP78 (an UPR initiation marker) using porcine-specific siRNAs (#909, #693, and #1570) on oocyte maturation. Among all siRNAs, siRNA #693 significantly reduced the protein levels of UPR marker proteins (BIP/GRP78, ATF4, and P90ATF6) in porcine COCs observed by Western blotting and immunofluorescence analysis. We also observed that the reduction of BIP/GRP78 levels by siRNA#693 significantly inhibited the meiotic maturation of oocytes (siRNA #693: $32.5{\pm}10.1%$ vs control: $77.8{\pm}5.3%$). In addition, we also checked the effect of ER-stress inhibitors, tauroursodeoxycholic acid (TUDCA, $200{\mu}M$) and melatonin ($0.1{\mu}M$), in BIP/GRP78-knockdown oocytes. TUDCA and melatonin treatment could restore the expression levels of ER-stress marker proteins (BIP/GRP78, $p-eIF2{\alpha}$, $eIF2{\alpha}$, ATF4, and P90ATF6) in siRNA #693-transfected matured COCs. In conclusion, these results demonstrated that BIP/GRP78-mediated regulation of UPR signaling and ER stress plays an important role in in vitro maturation of porcine oocytes.

• Journal of Life Science
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• v.23 no.8
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• pp.1050-1056
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• 2013

Streptococcus pneumoniae is the leading cause of community-acquired pneumonia throughout the world. The bacteria invade through lung tissue and cause sepsis, shock, and serious sequelae, including rheumatic fever and acute glomerulonephritis. However, the molecular mechanism associated with pneumonia's penetration of lung tissue and invasion of the blood stream are still unclear. We attempted to investigate the host cell response at protein levels to S. pneumoniae D39 invasion using human lung cancer epithelial cells, A549. Streptococcus pneumoniae D39 began to change the morphology of A549 cells to become round with filopodia at 2 hours post-infection. A549 cell proteins obtained at each infection time point were separated by SDS-PAGE and analyzed using MALDI-TOF. We identified several endoplasmic reticulum (ER) resident proteins such as Grp94 and Grp78 and mitochondrial proteins such as ATP synthase and Hsp60 that increased after S. pneumoniae D39 infection. Cytosolic Hsc70 and Hsp90 were, however, identified to decrease. These proteins were also confirmed by Western blot analysis. The identified ER resident proteins were known to be induced during ER stress signaling. These/ data, therefore, suggest that S. pneumoniae D39 infection may induce ER stress.

• Laboraroty Animal Research
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• v.34 no.4
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• pp.288-294
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• 2018

A few clues about correlation between endoplasmic reticulum (ER) stress and mulberry (Morus alba) leaves were investigated in only the experimental autoimmune myocarditis and streptozotocin-induced diabetes. To investigate whether a novel extract of mulberry leaves fermented with Cordyceps militaris (EMfC) could suppress ER in fatty liver, alterations in the key parameters for ER stress response were measured in high fat diet (HFD)-induced obese C57L/6 mice treated with EMfC for 12 weeks. The area of adipocytes in the liver section were significantly decreased in the HFD+EMfC treated group as compared to the HFD+Vehicle treated group, while their level was higher in HFD+Vehicle treated group than No treated group. The level of the eukaryotic initiation factor 2 alpha ($eIF2{\alpha}$) and inositol-requiring enzyme 1 beta ($IRE1{\alpha}$) phosphorylation and CCAAT-enhancer-binding protein homologous protein (CHOP) expression were remarkably enhanced in the HFD+Vehicle treated group. However, their levels were restored in the HFD+EMfC treated group, although some differences were detected in the decrease rate. Similar recovery was observed on the ER stress-induced apoptosis. The level of Caspase-3, Bcl-2 and Bax were decreased in the HFD+EMfC and HFD+orlistat (OT) treated group compared to the HFD+Vehicle treated group. The results of the present study therefore provide first evidence that EMfC with the anti-obesity effects can be suppressed ER stress and ER stress-induced apoptosis in the hepatic steatosis of HFD-induced obesity model.

• Journal of Physiology & Pathology in Korean Medicine
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• v.24 no.6
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• pp.1004-1011
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• 2010

This study was designed to evaluate the protective effects of Dodamtanggami-bang (DDTG) on tunicamycin induced cell death by ER stress in C6 glial cells. Cell viability was measured by MTT assay and LDH release. Apoptosis was determined by caspase activity and flow cytometry in C6 glial cells. Expression of ER stress mediators including, GRP78 and CHOP proteins were measured by Western blot analysis. Tunicamycin induced the apoptosis of C6 glial cells, which was characterized as nucleic acid and caspase-3 activation, PARP cleavage, and sub-G0/G1 fraction of cell cycle increase. However, pretreatment with DDTG protected C6 glial cells from tunicamycin. Treatment with tunicamycin resulted in the increased the expression of GRP78 and CHOP protein and produced ROS generation. However, pretreatment with DDTG inhibited the ER stress pathway, including increase of the expression of GRP78, CHOP proteins in C6 glial cells treated with tunicamycin. Taken together, these data suggest that DDTG is able to protect C6 glial cells from tunicamycin with marked inhibition of ER stress.

• Animal cells and systems
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• v.10 no.3
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• pp.115-120
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• 2006

Parkinson's disease (PD) is a neurodegenerative disease caused by selective degeneration of dopaminergic neurons in the substantia nigra. Mutations in ${\alpha}$-synuclein have been causally linked to the pathogenesis of hereditary PD. In addition, it is a major component of Lewy body found in the brains of sporadic cases as well. In the present study, we examined whether overexpression of wild type or PD-related mutant ${\alpha}$-synuclein induces unfolded protein response (UPR) and triggers the known signaling pathway of the resulting endoplasmic reticulum (ER) stress in SH-SY5Y cells. Overexpression of wild type, A30P, and A53T ${\alpha}$-synuclein all induced XBP-1 mRNA splicing, one of the late stage UPR events. However, activation of ER membrane kinases and upregulation of ER or cytoplsmic chaperones were not detected when ${\alpha}$-synuclein was overexpressed. However, basal level of cytoplsmic calcium was elevated in ${\alpha}$-synuclein-expressing cells. Our observation suggests that overexpression of ${\alpha}$-synuclein induces UPR independent of the known ER membrane kinase-mediated signaling pathway and induces ER stress by disturbing calcium homeostasis.

• BMB Reports
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• v.45 no.8
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• pp.482-487
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• 2012

To identify the novel inhibitors of endoplasmic reticulum stress-induced cell death, we performed a high throughput assay with a chemical library containing a total of 3,280 bioactive small molecules. Cyclosporine A and bromocriptine were identified as potent inhibitors of thapsigargiin-induced cell death (cut-off at $4{\sigma}$ standard score). However, U74389G, the potent inhibitor of lipid peroxidation had lower activity in inhibiting cell death. The inhibition effect of cyclosporine A and bromocriptine was specific for only thapsigargin-induced cell death. The mechanism of inhibition by these compounds was identified as modification of the expression of glucose regulated protein-78 (GRP-78/Bip) and inhibition of phosphorylation of p38 mitogen activated protein kinase (MAPK). However, these compounds did not inhibit the same events triggered by tunicamycin, which was in agreement with the cell survival data. We suggest that the induction of protective unfolded protein response by these compounds confers resistance to cell death. In summary, we identified compounds that may provide insights on cell death mechanisms stimulated by ER stress.

• Molecules and Cells
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• v.40 no.1
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• pp.66-72
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• 2017

Pathological hypertrophy of the heart is closely associated with endoplasmic reticulum stress (ERS), leading to maladaptations such as myocardial fibrosis, induction of apoptosis, and cardiac dysfunctions. Salubrinal is a known selective inhibitor of protein phosphatase 1 (PP1) complex involving dephosphorylation of phospho-eukaryotic translation initiation factor 2 subunit $(p-eIF2)-{\alpha}$, the key signaling process in the ERS pathway. In this study, the effects of salubrinal were examined on cardiac hypertrophy using the mouse model of transverse aortic constriction (TAC) and cell model of neonatal rat ventricular myocytes (NRVMs). Treatment of TAC-induced mice with salubrinal ($0.5mg{\cdot}kg^{-1}{\cdot}day^{-1}$) alleviated cardiac hypertrophy and tissue fibrosis. Salubrinal also alleviated hypertrophic growth in endothelin 1 (ET1)-treated NRVMs. Therefore, the present results suggest that salubrinal may be a potentially efficacious drug for treating pathological cardiac remodeling.

• Molecules and Cells
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• v.43 no.1
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• pp.48-57
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• 2020

The microalga Chlamydomonas reinhardtii accumulates triacylglycerols (TAGs) in lipid droplets under stress conditions, such as nitrogen starvation. TAG biosynthesis occurs mainly at the endoplasmic reticulum (ER) and requires fatty acid (FA) substrates supplied from chloroplasts. How FAs are transferred from chloroplast to ER in microalgae was unknown. We previously reported that an Arabidopsis thaliana ATP-binding cassette (ABC) transporter, AtABCA9, facilitates FA transport at the ER during seed development. Here we identified a gene homologous to AtABCA9 in the C. reinhardtii genome, which we named CrABCA2. Under nitrogen deprivation conditions, CrABCA2 expression was upregulated, and the CrABCA2 protein level also increased. CrABCA2 knockdown lines accumulated less TAGs and CrABCA2 overexpression lines accumulated more TAGs than their untransformed parental lines. Transmission electron microscopy showed that CrABCA2 was localized in swollen ER. These results suggest that CrABCA2 transports substrates for TAG biosynthesis to the ER during nitrogen starvation. Our study provides a potential tool for increasing lipid production in microalgae.

• Toxicological Research
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• v.24 no.2
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• pp.129-135
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• 2008

The results of recent studies indicate that high levels of free fatty acids(FFAs) and adipokines may be the main causes of non-alcoholic liver disease; however, the molecular mechanism that links FFAs to lipotoxicity remains unclear. In the present study, we treated HepG2 cells with FFA(either palmitate or oleate) to investigate the mechanisms involved in lipotoxicity in the liver cells. We also treated cells with palmitate in the presence of a chemical chaperone, 4-phenylbutyric acid(PBA), to confirm the involvement of ER stress in lipotoxicity. Palmitate significantly induced cytotoxicity in dose- and time-dependent manners. Apoptosis was also significantly induced by palmitate as measured by caspase-3 activity and DAPI staining. Palmitate led to increased expressions of the spliced form of X-box-protein(Xbp)-1 mRNA and C/EBP homologous transcription factor(CHOP) protein, suggesting activation of the unfolded-protein response. PBA co-incubation significantly attenuated apoptosis induced by palmitate. The above data demonstrate that high levels of palmitate induce apoptosis via the mediation of ER stress in the liver cells and that chemical chaperones act to modulate ER stress and accompanying apoptosis.