• Asian Pacific Journal of Cancer Prevention
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• v.15 no.3
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• pp.1099-1104
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• 2014

Aims and Background: Ginsenoside Rh2, which exerts the potent anticancer action both in vitro and in vivo, is one of the most well characterized ginsenosides extracted from ginseng. Although its effects on cancer are significant, the underlying mechanisms remain unknown. In this study, we sought to elucidate possible links between ginsenoside Rh2 and phosphoglucose isomerase/autocrine motility factor (PGI/AMF). Methods: $KG1{\alpha}$, a leukemia cell line highly expressing PGI/AMF was assessed by western blot analysis and reverse transcription- PCR (RT-PCR) assay after transfection of a small interfering (si)-RNA to silence PGI/AMF. The effect of PGI/AMF on proliferation was measured by typan blue assay and antibody array. A cell counting kit (CCK)-8 and flow cytometry (FCM) were adopted to investigate the effects of Rh2 on PGI/AMF. The relationships between PGI/AMF and Rh2 associated with Akt, mTOR, Raptor, Rag were detected by western blot analysis. Results: KG1${\alpha}$ cells expressed PGI/AMF and its down-regulation significantly inhibited proliferation. The antibody array indicated that the probable mechanism was reduced expression of PARP, State1, SAPK/JNK and Erk1/2, while those of PRAS40 and p38 were up-regulated. Silencing of PGI/AMF enhanced the sensibility of $KG1{\alpha}$ to Rh2 by suppressing the expression of mTOR, Raptor and Akt. Conclusion: These results suggested that ginsenoside Rh2 suppressed the proliferation of $KG1{\alpha}$, the same as down-regulation of PGI/AMF. Down-regulation of PGI/AMF enhanced the pharmacological effects of ginsenoside Rh2 on KG1${\alpha}$ by reducing Akt/mTOR signaling.

• Biomolecules & Therapeutics
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• v.30 no.4
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• pp.348-359
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• 2022

Gastric adenocarcinoma is among the top causes of cancer-related death and is one of the most commonly diagnosed carcinomas worldwide. Benzyl isothiocyanate (BITC) has been reported to inhibit the gastric cancer metastasis. In our previous study, BITC induced apoptosis in AGS cells. The purpose of the present study was to investigate the effect of BITC on autophagy mechanism in AGS cells. First, the AGS cells were treated with 5, 10, or 15 μM BITC for 24 h, followed by an analysis of the autophagy mechanism. The expression level of autophagy proteins involved in different steps of autophagy, such as LC3B, p62/SQSTM1, Atg5-Atg12, Beclin1, p-mTOR/mTOR ratio, and class III PI3K was measured in the BITC-treated cells. Lysosomal function was investigated using cathepsin activity and Bafilomycin A1, an autophagy degradation stage inhibitor. Methods including qPCR, western blotting, and immunocytochemistry were employed to detect the protein expression levels. Acridine orange staining and omnicathepsin assay were conducted to analyze the lysosomal function. siRNA transfection was performed to knock down the LC3B gene. BITC reduced the level of autophagy protein such as Beclin 1, class III PI3K, and Atg5-Atg12. BITC also induced lysosomal dysfunction which was shown as reducing cathepsin activity, protein level of cathepsin, and enlargement of acidic vesicle. Overall, the results showed that the BITC-induced AGS cell death mechanism also comprises the inhibition of the cytoprotective autophagy at both initiation and degradation steps.

• Journal of Life Science
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• v.23 no.5
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• pp.689-697
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• 2013

In the United States, about 40,000 new cases of oral cancer are diagnosed each year and nearly 7,800 patients died from it in 2012. Omega-3 polyunsaturated fatty acids have been found to have anticancer effects in a variety of cancer cell lines and animal models, but their effect in oral cancer remains unclear. This study was designed to examine the effect of docosahexaenoic acid (DHA, a kind of omega-3 fatty acid) on oral cancer cells and the molecular mechanism of its action. We found that exposure of squamous cell carcinoma-4 (SCC-4) and squamous cell carcinoma-9 (SCC-9) human oral cancer cells to DHA induced growth inhibition in a dose- and time-dependent manner. Meanwhile, in addition to the elevated levels of apoptotic markers, such as cleaved PARP, subG1 portion and TUNEL-positive nuclei, DHA led to autophagic vesicle formation and an increase in autophagic flux, indicating the involvement of both apoptosis and autophagy in the inhibitory effects of DHA on oral cancer cells. Further experiments revealed that the apoptosis and autophagy induced by DHA were linked to inhibition of mammalian target of rapamycin (mTOR) signaling by AKT inhibition and AMP-activated protein kinase (AMPK) activation in SCC-9 cells. Together, our results suggest that DHA induces apoptosis- and autophagy-associated cell death through the AMPK/AKT/mTOR signaling pathway in oral cancer cells. Thus, utilization of omega-3 fatty acids may represent a promising therapeutic approach for chemoprevention and treatment of human oral cancer.

• Journal of Applied Biological Chemistry
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• v.58 no.3
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• pp.219-226
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• 2015

The anticancer activity of a methanolic extract from lemon leaves (MLL) was assessed in MCF-7-SC human breast cancer stem cells. MLL induced apoptosis in MCF-7-SC, as evidenced by increased apoptotic body formation, sub-G1 cell population, annexin V-positive cells, Bax/Bcl-2 ratio, as well as proteolytic activation of caspase-9 and caspase-3, and degradation of poly (ADP-ribose) polymerase (PARP) protein. Concomitantly, MLL induced the formation of acidic vesicular organelles, increased LC3-II accumulation, and reduced the activation of Akt, mTOR, and p70S6K, suggesting that MLL initiates an autophagic progression in MCF-7-SC via the Akt/mTOR pathway. Epithelial-mesenchymal transition (EMT), a critical step in the acquisition of the metastatic state, is an attractive target for therapeutic interventions directed against tumor metastasis. At low concentrations, MLL induced anti-metastatic effects on MCF-7-SC by inhibiting the EMT process. Exposure to MLL also led to an increase in the epithelial marker E-cadherin, but decreased protein levels of the mesenchymal markers Snail and Slug. Collectively, this study provides evidence that lemon leaves possess cytotoxicity and anti-metastatic properties. Therefore, MLL may prove to be beneficial as a medicinal plant for alternative novel anticancer drugs and nutraceutical products.

• Biomolecules & Therapeutics
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• v.31 no.6
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• pp.599-610
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• 2023

According to recent evidence, ferroptosis is a major cell death mechanism in the pathogenesis of kidney injury and fibrosis. Despite the renoprotective effects of classical ferroptosis inhibitors, therapeutic approaches targeting kidney ferroptosis remain limited. In this study, we assessed the renoprotective effects of melatonin and zileuton as a novel therapeutic strategy against ferroptosis-mediated kidney injury and fibrosis. First, we identified RSL3-induced ferroptosis in renal tubular epithelial HK-2 and HKC-8 cells. Lipid peroxidation and cell death induced by RSL3 were synergistically mitigated by the combination of melatonin and zileuton. Combination treatment significantly downregulated the expression of ferroptosis-associated proteins, 4-HNE and HO-1, and upregulated the expression of GPX4. The expression levels of p-AKT and p-mTOR also increased, in addition to that of NRF2 in renal tubular epithelial cells. When melatonin (20 mg/kg) and zileuton (20 mg/kg) were administered to a unilateral ureteral obstruction (UUO) mouse model, the combination significantly reduced tubular injury and fibrosis by decreasing the expression of profibrotic markers, such as α-SMA and fibronectin. More importantly, the combination ameliorated the increase in 4-HNE levels and decreased GPX4 expression in UUO mice. Overall, the combination of melatonin and zileuton was found to effectively ameliorate ferroptosis-related kidney injury by upregulating the AKT/mTOR/ NRF2 signaling pathway, suggesting a promising therapeutic strategy for protection against ferroptosis-mediated kidney injury and fibrosis.

• Biomolecules & Therapeutics
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• v.25 no.6
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• pp.609-617
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• 2017

Pancreatic cancer is one of the most lethal and aggressive cancers in the world. However, no effective treatment is currently available for pancreatic cancer. The objective of this study was to determine the anti-pancreatic cancer effect of ${\alpha}$-mangostin (${\alpha}M$) and ${\gamma}$-mangostin (${\gamma}M$) extracted from the pericarp of Garcinia mangostana L.. Both ${\alpha}$M and ${\gamma}M$ reduced the viability of pancreatic cancer cells MIA PaCa-2 and PANC-1 in a dose-dependent manner. These compounds induced apoptosis by increasing c-PARP and c-Caspase 3 levels. They also induced autophagy by increasing levels of microtubule-associated protein 1A/1B light chain 3B (LC3II) in both cell lines while decreasing sequestosome 1 (p62) in MIA PaCa-2. Both ${\alpha}$M and ${\gamma}M$ induced autophagy through increasing phosphorylation levels of AMP-activated protein kinase (p-AMPK) and p38-mitogen activated protein kinase (p-p38) while decreasing phosphorylation level of mammalian target of rapamycin complex 1 (p-mTOR). Of various microRNAs (miRNA), miR-18a was found to be a putative regulatory miRNA for autophagy induced by ${\alpha}$M or ${\gamma}M$. In combination with gemcitabine, a compound frequently used in pancreatic cancer treatment, ${\alpha}$M and ${\gamma}M$ showed synergistic anti-cancer effects in MIA PaCa-2. Collectively, these results suggest that ${\alpha}$M and ${\gamma}M$ can induce apoptosis and autophagy in pancreatic cancer cells and that their anti-cancer effect is likely to be associated with miR-18a. In conclusion, ${\alpha}$M and ${\gamma}M$ might be used as a potential new therapy for pancreatic cancer.

• Journal of Food Hygiene and Safety
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• v.31 no.4
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• pp.294-298
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• 2016

Natural products are attractive as the source of new drug development. Especially, numerous unknown marine bioresources are an object of attention because the ocean occupies three fourth of the earth. Survival of marine bioresources in extreme environment may induce the production of biological active compounds. As previous study, we examined over 40 specimens of marine sponges collected from Micronesia and screened their anti-proliferative activities in various cancer cell lines. Among them, we investigated Coscinoderma sp.'s activity and mechanism in human colon carcinoma HCT116 and RKO cells. Furthermore, we also used the p53-knockout of HCT116 cells and the p53 loss of RKO cells for elucidating the role of p53. Coscinoderma sp. inhibited cellular viability independently of the p53 status. Therefore, we compared the expression level of cell death-related proteins by Coscinoderma sp. in HCT16 and in HCT116 p53KO cells. Coscinoderma sp. increased p53 level and NOXA levels and induced apoptosis under the condition of p53 existence. On the other hand, Coscinoderma sp. increased p21 and mTOR levels in HCT116 p53KO cells. These results suggest that Coscinoderma sp. induced anti-proliferation effect through different pathway depending on p53 status.

• Journal of Ginseng Research
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• v.44 no.1
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• pp.67-78
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• 2020

Background: Gintonin (GT), a novel ginseng-derived exogenous ligand of lysophosphatidic acid (LPA) receptors, has been shown to induce cell proliferation and migration in the hippocampus, regulate calcium-dependent ion channels in the astrocytes, and reduce β-amyloid plaque in the brain. However, whether GT influences autophagy in cortical astrocytes is not yet investigated. Methods: We examined the effect of GT on autophagy in primary cortical astrocytes using immunoblot and immunocytochemistry assays. Suppression of specific proteins was performed via siRNA. LC3 puncta was determined using confocal microscopy. Results: GT strongly upregulated autophagy marker LC3 by a concentration- as well as time-dependent manner via G protein-coupled LPA receptors. GT-induced autophagy was further confirmed by the formation of LC3 puncta. Interestingly, on pretreatment with an mammalian target of rapamycin (mTOR) inhibitor, rapamycin, GT further enhanced LC3-II and LC3 puncta expression. However, GT-induced autophagy was significantly attenuated by inhibition of autophagy by 3-methyladenine and knockdown Beclin-1, Atg5, and Atg7 gene expression. Importantly, when pretreated with a lysosomotropic agent, E-64d/peps A or bafilomycin A1, GT significantly increased the levels of LC3-II along with the formation of LC3 puncta. In addition, GT treatment enhanced autophagic flux, which led to an increase in lysosome-associated membrane protein 1 and degradation of ubiquitinated p62/SQSTM1. Conclusion: GT induces autophagy via mTOR-mediated pathway and elevates autophagic flux. This study demonstrates that GT can be used as an autophagy-inducing agent in cortical astrocytes.

• Journal of Convergence for Information Technology
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• v.11 no.2
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• pp.201-210
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• 2021

This study was carried out to investigate the Fagopyrum esculentum (F. esculentum) extracts and vitexin are as the results of microarray, cell proliferation, cell wound recovery, cell cycle, microphage pattern and protein analysis for damage improvement caused by UVB-induced damage. Microarray results showed that UVB-induced increase in DRAM1, Atg2a and Atg13 genes was reduced in F. esculentum ethanol extract and vitexin. Cell proliferation, wound repair, cell cycle, and microphage patterns were improved in F. esculentum ethanol extract and vitexin, while buckwheat ethanol extract and vitexin decreased in both DRAM1, Beclin-1, and LC3 I/II in the vitexin treatment group and p-mTOR and survivin were all increased in protein analysis. It is thought that it can recover to normal and control autophagy, one of the causes of cell aging caused by UVB, to inhibit and regenerate cell death. F. esculentum ethanol extract and vitexin can be used as a functional cosmetic ingredient.

• Molecules and Cells
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• v.46 no.4
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• pp.245-255
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• 2023

This study aimed to exploring the pathophysiological mechanism of 7α,25-dihydroxycholesterol (7α,25-DHC) in osteoarthritis (OA) pathogenesis. 7α,25-DHC accelerated the proteoglycan loss in ex vivo organ-cultured articular cartilage explant. It was mediated by the decreasing extracellular matrix major components, including aggrecan and type II collagen, and the increasing expression and activation of degenerative enzymes, including matrix metalloproteinase (MMP)-3 and -13, in chondrocytes cultured with 7α,25-DHC. Furthermore, 7α,25-DHC promoted caspase-dependent chondrocyte death via extrinsic and intrinsic pathways of apoptosis. Moreover, 7α,25-DHC upregulated the expression of inflammatory factors, including inducible nitric oxide synthase, cyclooxygenase-2, nitric oxide, and prostaglandin E₂, via the production of reactive oxygen species via increase of oxidative stress in chondrocytes. In addition, 7α,25-DHC upregulated the expression of autophagy biomarkers, including beclin-1 and microtubule-associated protein 1A/1B-light chain 3 via the modulation of p53-Akt-mTOR axis in chondrocytes. The expression of CYP7B1, caspase-3, and beclin-1 was elevated in the degenerative articular cartilage of mouse knee joint with OA. Taken together, our findings suggest that 7α,25-DHC is a pathophysiological risk factor of OA pathogenesis that is mediated a chondrocyte death via oxiapoptophagy, which is a mixed mode of apoptosis, oxidative stress, and autophagy.