• Maxillofacial Plastic and Reconstructive Surgery
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• 제35권3호
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• pp.143-154
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• 2013

Purpose: This study was to find efficacy of integrin alpha2 (${\alpha}_2$) and epidermal growth factor receptor (EGFR) as tumor marker of oral squamous cell carcinoma (SCC) and clarify the selective cell death effect of anti-integrin ${\alpha}_2$ and anti-EGFR on SCC cells, additionally testify conjugated gold nanoparticles (GNP) with air plasma for selective cell death of oral SCC. Methods: Expression of integrin ${\alpha}_2$, EGFR on human SCC cells (SCC25) were examined by western blot. SCC25 cells were treated with anti-integrin ${\alpha}_2$, anti-EGFR and analysed by Hemacolor staining, immunoflorescence staining, FACS flow cytometry. Conjugated GNP with integrin ${\alpha}_2$, EGFR antibody were treated by air plasma on SCC cells. Results: Integrin ${\alpha}_2$ and EGFR were over-expressed on SCC25 cells than normal lung WI-38 cells. The cell viability rate of SCC25 cells treated with anti-integrin ${\alpha}_2$, anti-EGFR was lower than WI-38 cells. The concentration changes of nucleus, releasing cytochrome c and apoptosis inducing factor (AIF) from mitochondria to cytosol were observed. The changes of proteins related with apoptosis were observed. Increase of bax, bcl-xL, activation of caspase-3, -7, -9, and fragmentation of PARP, DFF45 and decrease of lamin A/C in SCC25 cells were observed. In FACS, increase of sub-$G_1$ and S phase was observed. Cell cycle related proteins, Such as cyclin D1, cyclin dependent kinase (CDK) 4, cyclin A, cyclin E, CDK 2, p27 were decreased. After SCC25 cells treated with conjugatged GNP-Integrin ${\alpha}_2$, GNP-EGFR, additionally air plasma, the cell death rate was significantly increased. Conclusion: Integrin ${\alpha}_2$, EGFR were over-expressed in oral SCC cells. Anti-integrin ${\alpha}_2$, anti-EGFR in SCC25 cells induced apoptosis selectively. When GNP-anti integrin ${\alpha}_2$, GNP-anti EGFR were treated with air plasma on SCC25 cells, cancer cells were died more selectively. GNP-anti integrin ${\alpha}_2$, GNP-anti EGFR with air plasma could be treatment choice of oral SCC.

• Tuberculosis and Respiratory Diseases
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• 제82권2호
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• pp.133-142
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• 2019

Background: Idiopathic pulmonary fibrosis involves irreversible alveolar destruction. Although alveolar epithelial type II cells are key functional participants within the lung parenchyma, how epithelial cells are affected upon bleomycin (BLM) exposure remains unknown. In this study, we determined whether BLM could induce cell cycle arrest via regulation of Schlafen (SLFN) family genes, a group of cell cycle regulators known to mediate growth-inhibitory responses and apoptosis in alveolar epithelial type II cells. Methods: Mouse AE II cell line MLE-12 were exposed to $1-10{\mu}g/mL$ BLM and $0.01-100{\mu}M$ baicalein (Bai), a G1/G2 cell cycle inhibitor, for 24 hours. Cell viability and levels of pro-inflammatory cytokines were analyzed by MTT and enzyme-linked immunosorbent assay, respectively. Apoptosis-related gene expression was evaluated by quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR). Cellular morphology was determined after DAPI and Hoechst 33258 staining. To verify cell cycle arrest, propidium iodide (PI) staining was performed for MLE-12 after exposure to BLM. Results: BLM decreased the proliferation of MLE-12 cells. However, it significantly increased expression levels of interleukin 6, tumor necrosis factor ${\alpha}$, and transforming growth factor ${\beta}1$. Based on Hoechst 33258 staining, BLM induced condensation of nuclear and fragmentation. Based on DAPI and PI staining, BLM significantly increased the size of nuclei and induced G2/M phase cell cycle arrest. Results of qRT-PCR analysis revealed that BLM increased mRNA levels of BAX but decreased those of Bcl2. In addition, BLM/Bai increased mRNA levels of p53, p21, SLFN1, 2, 4 of Schlafen family. Conclusion: BLM exposure affects pulmonary epithelial type II cells, resulting in decreased proliferation possibly through apoptotic and cell cycle arrest associated signaling.

• Journal of Ginseng Research
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• 제41권3호
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• pp.247-256
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• 2017

Background: KG-135, a standardized formulation enriched with Rk1, Rg3, and Rg5 ginsenosides, has been shown to inhibit various types of cancer cells; however, the underlying mechanisms are not fully understood. In this study, we explored its effects in A549 human lung cancer cells to investigate the induction of Forkhead Class box O3a (FOXO3a) and autophagy. Methods: Cell viability was determined by sulforhodamine B staining. Apoptosis and cell cycle distribution were analyzed using flow cytometry. The changes of protein levels were determined using Western blot analysis. Autophagy induction was monitored by the formation of acidic vesicular organelles stained with acridine orange. Results: KG-135 effectively arrested the cells in G1 phase with limited apoptosis. Accordingly, a decrease of cyclin-dependent kinase-4, cyclin-dependent kinase-6, cyclin D1, and phospho-retinoblastoma protein, and an increase of p27 and p18 proteins were observed. Intriguingly, KG-135 increased the tumor suppressor FOXO3a and induced the accumulation of autophagy hallmark LC3-II and acidic vesicular organelles without an increase of the upstream marker Beclin-1. Unconventionally, the autophagy adaptor protein p62 (sequestosome 1) was increased rather than decreased. Blockade of autophagy by hydroxychloroquine dramatically potentiated KG-135-induced FOXO3a and its downstream (FasL) ligand accompanied by the cleavage of caspase-8. Meanwhile, the decrease of Bcl-2 and survivin, as well as the cleavage of caspase-9, were also drastically enhanced, resulting in massive apoptosis. Conclusion: Besides arresting the cells in G1 phase, KG-135 increased FOXO3a and induced an unconventional autophagy in A549 cells. Both the KG-135-activated extrinsic FOXO3a/FasL/caspase-8 and intrinsic caspase-9 apoptotic pathways were potentiated by blockade of autophagy. Combination of KG-135 and autophagy inhibitor may be a novel strategy as an integrative treatment for cancers.

• Asian-Australasian Journal of Animal Sciences
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• 제27권5호
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• pp.635-647
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• 2014

Unfertilized oocytes age inevitably after ovulation, which limits their fertilizable life span and embryonic development. Rapamycin affects mammalian target of rapamycin (mTOR) expression and cytoskeleton reorganization during oocyte meiotic maturation. The goal of this study was to examine the effects of rapamycin treatment on aged porcine oocytes and their in vitro development. Rapamycin treatment of aged oocytes for 24 h (68 h in vitro maturation [IVM]; $44h+10{\mu}M$ rapamycin/24 h, $47.52{\pm}5.68$) or control oocytes (44 h IVM; $42.14{\pm}4.40$) significantly increased the development rate and total cell number compared with untreated aged oocytes (68 h IVM, $22.04{\pm}5.68$) (p<0.05). Rapamycin treatment of aged IVM oocytes for 24 h also rescued aberrant spindle organization and chromosomal misalignment, blocked the decrease in the level of phosphorylated-p44/42 mitogen-activated protein kinase (MAPK), and increased the mRNA expression of cytoplasmic maturation factor genes (MOS, BMP15, GDF9, and CCNB1) compared with untreated, 24 h-aged IVM oocytes (p<0.05). Furthermore, rapamycin treatment of aged oocytes decreased reactive oxygen species (ROS) activity and DNA fragmentation (p<0.05), and downregulated the mRNA expression of mTOR compared with control or untreated aged oocytes. By contrast, rapamycin treatment of aged oocytes increased mitochondrial localization (p<0.05) and upregulated the mRNA expression of autophagy (BECN1, ATG7, MAP1LC3B, ATG12, GABARAP, and GABARAPL1), anti-apoptosis (BCL2L1 and BIRC5; p<0.05), and development (NANOG and SOX2; p<0.05) genes, but it did not affect the mRNA expression of pro-apoptosis genes (FAS and CASP3) compared with the control. This study demonstrates that rapamycin treatment can rescue the poor developmental capacity of aged porcine oocytes.

• 대한한의학방제학회지
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• 제24권3호
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• pp.163-174
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• 2016

Objectives : Rheum undulatum Linne and Glycyrriza uralensis Fischer are widely used herbal medicine. In this study, anti-oxidant and liver protective effects of R. undunlatum extract (RUE) and G. uralensis extract (GUE) were investigated in HepG2 cells, respectively. Oxidative stress and liver fibrosis were induced by arachidonic acid (AA) and iron, and CCl₄.Methods : MTT assay was assessed for cell viability, and immunoblotting analysis was performed to detect expression of apoptosis related proteins. In addition, reactive oxygen species (ROS) and mitochondrial dysfunction were measured. In vivo, BALB/c mouse were orally administrated with the aqueous extract of 10 mg/kg RUE and 100 mg/kg GUE for 3 days and then, injected with CCl₄ 0.5 ml/kg body weight to induce acute liver damage. Serum ALT level was measured, and histological change was observed in Harris's hematoxylin and eosin stainResults : RUE and GUE pre-treatment increased relative cell viability in concentration dependent manner and altered the expression levels of apoptosis-related proteins such as procaspase 3, PARP and Bcl-xL. RUE and GUE also inhibited the mitochondrial dysfunction and excessive reactive oxygen species (ROS) production induced by AA and iron. In addition, RUE and GUE activated liver kinase B1 (LKB1), by increasing phosphorylation. Moreover, RUE and GUE treatment decreased liver injuries induced by CCl₄, as evidenced by decreases in histological liver damage as well as serum alanine amino transferase (ALT) level.Conclusions : These data suggest that RUE and GUE has anti-oxidant and liver protective effects against AA and iron-induced oxidative stress and CCl₄-induced liver injury.

• Journal of Oral Medicine and Pain
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• 제36권3호
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• pp.147-160
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• 2011

Chios gum mastic (CGM)은 그리이스 키오스 섬에서만 자생하는 Pistiacia lentiscus L. var. Chia. 의 잎과 줄기로부터 추출되어진 식물성 수지이며, 과거 수세기 동안 지중해와 중동 지역 국가들에서 음식 첨가물과 위궤양, 십이지장궤양 등의 민간 치료약재로서 사용되어져 왔다. 정향나무에서 추출하는 페놀화합물인 eugenol은 zinc oxide eugenol의 구성성분으로 치과치료를 위해 많이 사용되고 있다. 본 연구는 사람혀편평세포암종세포(SCC25 cells)에서 천연물질인 CGM과 eugenol을 병용처리한 후 세포자멸사 효과가 있는지를 알아보기 위해 수행하였다. CGM과 eugenol의 병용처리가 단독처리에 비해서 효과적인 세포생존율 감소가 있는지 확인하기 위하여 MTT법을 시행하였고, 세포자멸사의 유도와 증가를 알기 위하여 Hoechst 염색법, TUNEL 염색법, DNA hypoploidy법을 사용하였다. 그리고 세포자멸사에 관계하는 단백질의 발현 변화와 세포내에서의 이동을 밝혀내기 위하여 Western blot 분석과 면역형광염색법을 수행하였다. 본 연구에서는 CGM과 eugenol이 병용처리된 SCC25 세포에서 핵의 농축, DNA분절, Bax의 증가와 Bcl-2의 감소, DNA양의 감소, cytochrome c의 세포질로의 유리, AIF와 DFF40 (CAD)의 핵으로의 이동, caspase-3, caspase-6, caspase-7, caspase-9, PARP, Lamin A/C 그리고 DFF45 (ICAD)의 활성화와 같은 다양한 세포자멸사 증거를 보였다. 반면에 CGM과 eugenol이 각각 단독 처리된 SCC25 세포에서는 세포자멸사 현상이 미미하였다. 24시간 동안 40 ${\mu}g$/ml의 CGM과 0.5 mM의 eugenol을 각기 단독처리 한 결과에서는 세포자멸사를 거의 유도하지 못했으나, 병용처리 한 결과에는 아주 탁월한 세포자멸사의 유도를 보였다. 그러므로 본 실험결과는 사람구강편평세포암종 환자들에게 CGM과 eugenol의 병용요법이 새로운 치료전략으로서의 가능성을 줄 수 있다고 생각한다.

• 생명과학회지
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• 제19권2호
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• pp.249-255
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• 2009

Esculetin, a coumarin compound, has been known to inhibit proliferation and induce apoptosis in several types of human cancer cells. However, the molecular mechanisms involved in esculetin-induced apoptosis are still uncharacterized in human leukemia cells. In this study, we have investigated whether esculetin exerts anti-proliferative and apoptotic effects on human leukemia U937 cells. It was found that esculetin could inhibit cell viability in a time-dependent manner, which was associated with the induction of apoptotic cell death such as increased populations of apoptotic- sub G1 phase. Apoptosis of U937 cells by esculetin was associated with an inhibition of Bcl-2/Bax binding activity, formation of tBid, down-regulation of X-linked inhibitor of apoptotic protein (XIAP) expression, and up-regulation of death receptor 4 (DR4) and FasL expression. Esculetin treatment also induced the degradation of ${\beta}$-catenin and DNA fragmentation factor 45/inhibitor of caspase-activated DNase (DFF45/ICAD). Furthermore, a caspase-3 specific inhibitor, z-DEVD-fmk, significantly inhibited sub-G1 phase DNA content, morphological changes and degradation of ${\beta}$-catenin and DEE45/ICAD. These results indicated that a key regulator in esculetin-induced apoptosis was caspase-3 in human leukemia U937 cells.

• Laboraroty Animal Research
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• 제34권4호
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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.

• Animal Bioscience
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• 제35권9호
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• pp.1340-1350
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• 2022

Objective: Luxi Black Head sheep (LBH) is the first crossbreed specialized for meat production and was developed by crossbreeding Black Head Dorper sheep (DP) and Small Tailed Han sheep (STH) in the farming areas of northern China. Research on the genomic variations and selection signatures of LBH caused by continuous artificial selection is of great significance for identifying the genetic mechanisms of important traits of sheep and for the continuous breeding of LBH. Methods: We explored the genetic relationships of LBH, DP, and several Mongolian sheep breeds by constructing phylogenetic tree, principal component analysis and linkage disequilibrium analysis. In addition, we analysed 29 whole genomes of sheep. The genome-wide selection signatures have been scanned with four methods: heterozygosity (H_P), fixation index (F_ST), cross-population extended haplotype homozygosity (XP-EHH) and the nucleotide diversity (𝜃_π) ratio. Results: The genetic relationships analysis showed that LBH appeared to be an independent cluster closer to DP. The candidate signatures of positive selection in sheep genome revealed candidate genes for developmental process (HoxA gene cluster, BCL2L11, TSHR), immunity (CXCL6, CXCL1, SKAP2, PTK6, MST1R), growth (PDGFD, FGF18, SRF, SOCS2), and reproduction (BCAS3, TRIM24, ASTL, FNDC3A). Moreover, two signalling pathways closely related to reproduction, the thyroid hormone signalling pathway and the oxytocin signalling pathway, were detected. Conclusion: The selective sweep analysis of LBH genome revealed candidate genes and signalling pathways associated with developmental process, immunity, growth, and reproduction. Our findings provide a valuable resource for sheep breeding and insight into the mechanisms of artificial selection.

• Biomolecules & Therapeutics
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• 제31권1호
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• pp.73-81
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• 2023

Sirtuins (SIRTs) belong to the nicotinamide adenine dinucleotide (NAD+)-dependent class III histone deacetylase family. They are key regulators of cellular and physiological processes, such as cell survival, senescence, differentiation, DNA damage and stress response, cellular metabolism, and aging. SIRTs also influence carcinogenesis, making them potential targets for anticancer therapeutic strategies. In this study, we investigated the anticancer properties and underlying molecular mechanisms of a novel SIRT1 inhibitor, MHY2251, in human colorectal cancer (CRC) cells. MHY2251 reduced the viability of various human CRC cell lines, especially those with wild-type TP53. MHY2251 inhibited SIRT1 activity and SIRT1/2 protein expression, while promoting p53 acetylation, which is a target of SIRT1 in HCT116 cells. MHY2251 treatment triggered apoptosis in HCT116 cells. It increased the percentage of late apoptotic cells and the sub-G1 fraction (as detected by flow cytometric analysis) and induced DNA fragmentation. In addition, MHY2251 upregulated the expression of FasL and Fas, altered the ratio of Bax/Bcl-2, downregulated the levels of pro-caspase-8, -9, and -3 proteins, and induced subsequent poly(ADP-ribose) polymerase cleavage. The induction of apoptosis by MHY2251 was related to the activation of the caspase cascade, which was significantly attenuated by pre-treatment with Z-VAD-FMK, a pan-caspase inhibitor. Furthermore, MHY2251 stimulated the phosphorylation of c-Jun N-terminal kinase (JNK), and MHY2251-triggered apoptosis was blocked by pre-treatment with SP600125, a JNK inhibitor. This finding indicated the specific involvement of JNK in MHY2251-induced apoptosis. MHY2251 shows considerable potential as a therapeutic agent for targeting human CRC via the inhibition of SIRT1 and activation of JNK/p53 pathway.