• Journal of Microbiology and Biotechnology
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• v.32 no.5
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• pp.645-656
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• 2022

Gossypol, a natural phenolic aldehyde present in cotton plants, was originally used as a means of contraception, but is currently being studied for its anti-proliferative and anti-metastatic effects on various cancers. However, the intracellular mechanism of action regarding the effects of gossypol on pancreatic cancer cells remains unclear. Here, we investigated the anti-cancer effects of gossypol on human pancreatic cancer cells (BxPC-3 and MIA PaCa-2). Cell counting kit-8 assays, annexin V/propidium iodide staining assays, and transmission electron microscopy showed that gossypol induced apoptotic cell death and apoptotic body formation in both cell lines. RNA sequencing analysis also showed that gossypol increased the mRNA levels of CCAAT/enhancer-binding protein homologous protein (CHOP) and activating transcription factor 3 (ATF3) in pancreatic cancer cell lines. In addition, gossypol facilitated the cleavage of caspase-3 via protein kinase RNA-like ER kinase (PERK), CHOP, and Bax/Bcl-2 upregulation in both cells, whereas the upregulation of ATF was limited to BxPC-3 cells. Finally, a three-dimensional culture experiment confirmed the successful suppression of cancer cell spheroids via gossypol treatment. Taken together, our data suggest that gossypol may trigger apoptosis in pancreatic cancer cells via the PERK-CHOP signaling pathway. These findings propose a promising therapeutic approach to pancreatic cancer treatment using gossypol.

• The Korean Journal of Physiology and Pharmacology
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• v.72 no.4
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• pp.517-528
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• 2021

Aamylin or islet amyloid polypeptide (IAPP) is a peptide synthesized and secreted with insulin by the pancreatic β-cells. A role for amylin in the pathogenesis of type 2 diabetes (T2D) by causing insulin resistance or inhibiting insulin synthesis and secretion has been suggested by in vitro and in vivo studies. These studies are consistent with the effect of endogenous amylin on pancreatic β-cells to modulate and/or restrain insulin secretion. Here, we reported the correlation between amylin and insulin in rat insulinoma inS-1e cells by treating 2-deoxy-ᴰ-glucose (2-DG) and/or mannose. Cell viability was not affected by 24 h treatment with 2-DG and/or mannose, but it was significantly decreased by 48 h treatment with 5 and 10 mm 2-DG. in the 24 h treatment, the synthesis of insulin in the cells and the secretion of insulin into the media showed a significant inverse association. in the 48-h treatment, amylin synthesis vs. the secretion and insulin synthesis vs. the secretion showed a significant inverse relation. The synthesis of amylin vs. insulin and the secretion of amylin vs. insulin showed a significant inverse relationship. The p-ERK, antioxidant enzymes (Cu/Zn-superoxide dismutase (SOD), Mn-SOD, and catalase), and endoplasmic reticulum (ER) stress markers (cleaved caspase-12, CHOP, p-SAPK/JNK, and BiP/GRP78) were significantly increased or decreased by the 24 h and 48 h treatments. These data suggest the relative correlation to the synthesis of amylin by cells vs. the secretion into the media, the synthesis of amylin vs. insulin, and the secretion of amylin vs. insulin under 2-DG and/or mannose in rat insulinoma INS-1E cells. Therefore, these results can provide primary data for the hypothesis that the amylin-insulin relationships may be involved with the human amylin toxicity in pancreatic beta cells.

• The Korean Journal of Physiology and Pharmacology
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• v.9 no.6
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• pp.341-346
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• 2005

The mechanism underlying oxidant-induced intracellular $Ca^{2+}$ ($[Ca^{2+}]_i$) increase was studied in cultured bovine aortic endothelial cells (BAECs) using fura-2 AM. In the presence of 2 mM extracellular $Ca^{2+}$, the application of DTBNP ($20{\mu}M$), a membrane-permeable oxidant, caused an increase in $[Ca^{2+}]_i$, and DTT (2 mM) as a reductant completely reversed the effect of DTBNP. The $[Ca^{2+}]_i$ increase induced by DTBNP was also observed in an extracellular $Ca^{2+}$-free/2 mM EGTA solution, indicating the release of $Ca^{2+}$ from intracellular store(s). After endoplasmic reticulum was depleted by an $IP_3$-generating agonist, ATP ($30{\mu}M$) or an ER $Ca^{2+}$ pump inhibitor, thapsigargin ($1{\mu}M$), DTBNP-stressed BAECs showed an increase of $[Ca^{2+}]_i$ in $Ca^{2+}$-free/2 mM EGTA solution. Ratio-differences before and after the application of DTBNP after pretreatment with ATP or thapsigargin were $0.42{\pm}0.15$ and $0.49{\pm}0.07$, respectively (n=7), which are significantly reduced, compared to the control value of $0.72{\pm}0.07$ in a $Ca^{2+}$-free/2 mM EGTA solution. After the protonophore CCCP ($10{\mu}M$) challenge to release mitochondrial $Ca^{2+}$, the similar result was obtained. Ratio-difference before and after the application of DTBNP after pretreatment with CCCP was $0.46{\pm}0.09$ (n=7). Simultaneous application of thapsigargin and CCCP completely abolished the DTBNP-induced $[Ca^{2+}]_i$ increase. The above results together indicate that the increase of $[Ca^{2+}]_i$ by DTBNP resulted from the release of $Ca^{2+}$ from both endoplasmic reticulum and mitochondria.

• Applied Microscopy
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• v.29 no.2
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• pp.149-162
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• 1999

For investigation of the activated carbon on the mouse kidney treated with Pb, the activated carbon (40 mg/kg) and Pb (30 mg/kg) were treated orally for three and six weeks, respectivelly, and observed by the electron microscope. On the glomerulus of the group with only Pb, the basal membrane thicked, projected, and the foot processes fused. On the proximal convoluted tubules, the number of microvilli were decreased and the number of vacuoles and lysosome increased on the cytoplasm. The mitochondria and endoplasmic reticulum were extended and ribosomes dropped from the ER. On the giomerulus of the group with Pb-activated carbon, the basal membrane and the foot processes were merely changed. On the proximal convoluted tubules, the shapes and number of microvilli were not changed and the number of vacuoles, microbodies, and lysosomes decreased. The shapes of mitochondria and endoplasmic reticulum observed almost similar with control group. Th at is, elongated mitochondria and attached ribosomes to endoplasmic reticulum. As result, the activated carbon has positive effect on reducing toxicity of lead in the mouse kidney in the view of electron microscope.

• Applied Microscopy
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• v.30 no.2
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• pp.165-172
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• 2000

To investigate the defensive effect of green tea against the lead toxicity, Sprague-Dewley rats (150 gm) were divided into 5 groups; the control group, the group treated with lead for 4 weeks (Group A-1), the group treated with lead and green tea for 4 weeks (Group A-2), the group treated with lead for 8 weeks (Group B-1), and the group treated with lead and green tea for 8 weeks (Group B-2). The lead acetate (500 ppm) was injected two times for one week into the abdomen and green tea solution (3 g/100 ml distilled water) offered freely. In the liver cell, The cristae of mitochondria were enlarged, the rough endoplasmic reticulum (rER) extended and many lysosome observed on the Group A- 1. The cristae of mitochondria were enlarged and many lysosome observed, but nucleus was normal on the Croup A-2. All of observed parts on the Group B-1 deeply wounded by lead than the group A-1. In the Group B-2, the mitochondria and rER were increased in number than the Group B-1 and the nucleus was observed normally.

• Molecules and Cells
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• v.37 no.9
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• pp.672-684
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• 2014

The exact causes of cell death in Parkinson's disease (PD) remain unknown despite extensive studies on PD.The identification of signaling and metabolic pathways involved in PD might provide insight into the molecular mechanisms underlying PD. The neurotoxin 1-methyl-4-phenylpyridinium ($MPP^+$) induces cellular changes characteristic of PD, and $MPP^+$-based models have been extensively used for PD studies. In this study, pathways that were significantly perturbed in $MPP^+$-treated human neuroblastoma SH-EP cells were identified from genome-wide gene expression data for five time points (1.5, 3, 9, 12, and 24 h) after treatment. The mitogen-activated protein kinase (MAPK) signaling pathway and endoplasmic reticulum (ER) protein processing pathway showed significant perturbation at all time points. Perturbation of each of these pathways resulted in the common outcome of upregulation of DNA-damage-inducible transcript 3 (DDIT3). Genes involved in ER protein processing pathway included ubiquitin ligase complex genes and ER-associated degradation (ERAD)-related genes. Additionally, overexpression of DDIT3 might induce oxidative stress via glutathione depletion as a result of overexpression of CHAC1. This study suggests that upregulation of DDIT3 caused by perturbation of the MAPK signaling pathway and ER protein processing pathway might play a key role in $MPP^+$-induced neuronal cell death. Moreover, the toxicity signal of $MPP^+$ resulting from mitochondrial dysfunction through inhibition of complex I of the electron transport chain might feed back to the mitochondria via ER stress. This positive feedback could contribute to amplification of the death signal induced by $MPP^+$.

• Animal cells and systems
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• v.2 no.1
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• pp.145-152
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• 1998

The spinning apparatus and production of secretory silk from silk gland of the black widow spider, Latrodectus mactans were studied with scanning and transmission electron microscopes. The silk glands were located in seven groups on the spinnerets including each pair of major and minor ampullate, 3 pairs of tubuliform, 1 pair of flagelliform, 2 pairs of aggregate, about 50 pairs of pyriform and over 250 pairs of aciniform glands, respect- ively. Each group of silk gland feeds silk into one of the three spinneret pairs. Secretory silk is synthesized from rough endoplasmic reticulum (rER) of glandular epithelial cells. The secretory silk is transported from toe rER into the secretory vacuoles which are grown up by fusion with the surrounding small vesicles including the secretory silk. The secretory vacuoles, which show a gradual increase in electron density with the process of maturity, are formed without involvement of the Golgi complex, suggesting that they do not play an important role in the processing of the secretory silk. The secretory silk products are released by the mechanism of apocrine secretion, losing part of their cytoplasm. Moreover, another type of silk precursor, possibly protein, appears as granular material, and is also discharged to the luminal cavity.

• BMB Reports
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• v.50 no.8
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• pp.411-416
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• 2017

The endoplasmic reticulum (ER) membrane protein complex subunit 6 (EMC6) is a novel human autophagy-related molecule. Here, using tissue microarray and immunohistochemistry, we report that EMC6 protein is lost or reduced in glandular cells of patients with gastric adenocarcinoma, compared to normal stomach mucosa. Overexpression of EMC6 in gastric cancer cells inhibited cell growth, migration, invasion, and induced apoptosis and cell cycle arrest at S-phase. Further investigation suggested that EMC6 overexpression in BGC823 human adenocarcinoma gastric cancer cells reduced tumorigenicity in a xenograft model, demonstrating that EMC6 has the characteristics of a tumor suppressor. This is the first study to show that EMC6 induces cell death in gastric cancer cells. The molecular mechanism of how EMC6 functions as a tumor suppressor needs to be further explored.

• BMB Reports
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• v.44 no.11
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• pp.735-740
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• 2011

Tunicamycin, an endoplasmic reticulum (ER) stress inducer, specifically inhibits N-glycosylation. The cyclic AMP (cAMP) response element-binding protein H (CREBH) was previously shown to be regulated by UPR-dependent proteolytic cleavage in the liver. On the other hand, the role of CREBH in other tissues is unknown. In the present study, tunicamycin increased the level of CREBH activation (cleavage) as well as mRNA expression in osteoblast cells. Adenoviral (Ad) overexpression of CREBH suppressed BMP2-induced expression of alkaline phosphatase (ALP) and osteocalcin (OC). Interestingly, the BMP2-induced OASIS (structurally similar to CREBH, a positive regulator of osteoblast differentiation) expression was also inhibited by CREBH overexpression. In addition, inhibition of CREBH expression using siRNA reversed the tunicamycin-suppressed ALP and OC expression. These results suggest that CREBH inhibited osteoblast differentiation via suppressing BMP2-induced ALP, OC and OASIS expression in mouse calvarial derived osteoblasts.

• BMB Reports
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• v.47 no.2
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• pp.69-79
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• 2014

$Ca^{2+}$ release from intracellular stores and influx from extracellular reservoir regulate a wide range of physiological functions including muscle contraction and rhythmic heartbeat. One of the most ubiquitous pathways involved in controlled $Ca^{2+}$ influx into cells is store-operated $Ca^{2+}$ entry (SOCE), which is activated by the reduction of $Ca^{2+}$ concentration in the lumen of endoplasmic or sarcoplasmic reticulum (ER/SR). Although SOCE is pronounced in non-excitable cells, accumulating evidences highlight its presence and important roles in skeletal muscle and heart. Recent discovery of STIM proteins as ER/SR $Ca^{2+}$ sensors and Orai proteins as $Ca^{2+}$ channel pore forming unit expedited the mechanistic understanding of this pathway. This review focuses on current advances of SOCE components, regulation and physiologic and pathophysiologic roles in muscles. The specific property and the dysfunction of this pathway in muscle diseases, and new directions for future research in this rapidly growing field are discussed.