• Journal of Ginseng Research
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• 제37권2호
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• pp.201-209
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• 2013

Ginsenoside, one of the active ingredients of Panax ginseng, has a variety of physiologic and pharmacologic effects. The purpose of this study was to explore the effects of ginsenoside Rd (G-Rd) on melastatin type transient receptor potential 7 (TRPM7) channels with respect to the proliferation and survival of AGS and MCF-7 cells (a gastric and a breast cancer cell line, respectively). AGS and MCF-7 cells were treated with different concentrations of G-Rd, and caspase-3 activities, mitochondrial depolarizations, and sub-G1 fractions were analyzed to determine if cell death occurred by apoptosis. In addition, human embryonic kidney (HEK) 293 cells overexpressing TRPM7 channels were used to confirm the role of TRPM7 channels. G-Rd inhibited the proliferation and survival of AGS and MCF-7 cells and enhanced caspase-3 activity, mitochondrial depolarization, and sub-G1 populations. In addition, G-Rd inhibited TRPM7-like currents in AGS and MCF-7 cells and in TRPM7 channel overexpressing HEK 293 cells, as determined by whole cell voltage-clamp recordings. Furthermore, TRPM7 overexpression in HEK 293 cells promoted G-Rd induced cell death. These findings suggest that G-Rd inhibits the proliferation and survival of gastric and breast cancer cells by inhibiting TRPM7 channel activity.

• Clinical and Experimental Pediatrics
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• 제51권10호
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• pp.1112-1117
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• 2008

목 적: 해마 절편 배양에서 산소-포도당 박탈(oxygen-glucose deprivation, OGD)에 의한 세포 사망과 신경 세포 사멸을 propidium iodide(PI) 섭취, Fluoro-Jade(FJ) 염색, TUNEL 염색, caspase-3 면역형광염색 방법으로 관찰하고자 하였다. 방 법: 생후 7일된 Sprague-Dawley 흰쥐의 해마를 MacIlwain chopper로 $350{\mu}m$ 두께의 절편으로 절단하였다. 해마 절편을 6-well plate의 insert 내의 반 유공(sem-porous) 막 위에서 membrane-interface technique으로 10일 동안 배양하였다. 배양된 해마 절편에 산소-포도당 박탈을 60분 동안 가한 후 재산소-재관류하에 기초 배양액에서 48시간 배양하였다. 재산소-재관류 동안 PI 섭취 형광 정도를 시간에 따라 형광 현미경으로 관찰하고 세포사망 백분율(percent cell death)을 측정하였다. 산소-포도당 박탈 직전과 24 시간 후에 해마 절편을 $15{\mu}m$ 두께로 냉동 절단 후 FJ 염색, TUNEL 염색, caspase-3 면역형광염색을 시행하여 세포 사망을 관찰하였다. 결과: OGD 후 PI 섭취 는 해마 절편의 CA1과 DG에 한정되어있었다. OGD 후 재산소-재관류 동안 6시간에서 48시간까지 PI 섭취 형광 강도는 시간이 증가함에 따라 증가하였다. 세포 사망 백분율은 CA1과 DG에서 모두 OGD 후 재산소-재관류 시간이 증가함에 따라 의미 있게 증가하였다(P<0.05). OGD 후 24시간에 세포 변성을 의미하는 많은 FJ 염색 양성 신경 세포 들이 CA1과 DG에서 관찰되었다. 고배율 confocal laser 현미경으로 관찰한 CA1에서의 신경 세포들 중 일부는 명확한 핵과 돌기를 가지고 있는 것을 보여 주었으며, 다른 신경 세포들은 핵의 분절화, 돌기의 손실 등을 보여 주었다. TUNEL 염색과 caspase-3 염색은 OGD 후 24시간에 CA1과 DA에서 TUNEL 양성 발현을 증가시키고 caspase-3 발현을 증가시켰다. 결 론: 해마 절편 배양에서 산소-포도당 박탈 에 의한 다수의 세포 사망을 관찰할 수 있었다. 사망한 세포 들은 주로 신경 세포의 caspase-3 활성화에 의해 매개된 사멸을 보였다.

• 대한약침학회지
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• 제17권2호
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• pp.7-17
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• 2014

Objectives: Condurango is widely used in various systems of complementary and alternative medicines (CAM) against oesophageal and stomach ailments including certain types of cancer. However, until now no systematic study has been conducted to verify its efficacy and dose with proper experimental support. Therefore, we examined if ethanolic extract of Condurango could ameliorate benzo[a]pyrene (BaP)-induced lung cancer in rats, in vivo to validate its use as traditional medicine. Methods: Fifteen male and 15 female Sprague-Dawley (SD) rats were treated with 0.28 mg/kg of Sweet Bee Venom (SBV) (high-dosage group) and the same numbers of male and female SD rats were treated with 0.2 mL/kg of normal saline (control group) for 13 weeks. We selected five male and five female SD rats from the high-dosage group and the same numbers of male and female SD rats from the control group, and we observed these rats for four weeks. We conducted body-weight measurements, ophthalmic examinations, urinalyses and hematology, biochemistry, histology tests. Results: A histological study revealed gradual progress in lung tissue-repair activity in Condurango-fed cancer-bearing rats, showing gradual tissue recovery after three months of drug administration. Condurango has the capacity to generate reactive oxygen species (ROS), which may contribute to a reduction in anti-oxidative activity and to an induction of oxidative stress-mediated cancer cell-death. Condurango-activated pro-apoptotic genes (Bax, caspase-3, caspase-9, p53, cytochrome-c, apaf-1, ICAD and PARP) and down-regulated antiapoptotic-Bcl-2 expression were noted both at mRNA and protein levels. Studies on caspase-3 activation and PARP cleavage by western blot analysis revealed that Condurango induced apoptosis through a caspase-3-dependent pathway. Conclusion: The anticancer efficacy of an ethanolic extract of Condurango for treating BaP-induced lung cancer in rats lends support for its use in various traditional systems of medicine.

• BMB Reports
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• 제38권6호
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• pp.709-716
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• 2005

Apoptosis and necrosis are distinguished by modality primarily. Here we show an apoptosis occurred instantly, induced by $300\;{\mu}M$ W-7 ((N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride), inhibitor of calmodulin), which demonstrated necrotic modality. As early as 30 min after W-7 addition, apoptotic (sub-diploid) peak could be detected by fluorescence-activated cell sorter (FACS), “DNA ladders” began to emerge also at this time point, activity of caspase-3 elevated obviously within this period. Absence of mitochondrial membrane potential (MMP) reduction and cytochrome c, AIF (apoptosis inducing factor) release, verified that this rapid apoptosis did not proceed through mitochondria pathway. Activation of caspase-12 and changes of other endoplasmic reticulum (ER) located proteins ascertained that ER pathway mediated this necrosis-like apoptosis. Our findings suggest that it is not credible to judge apoptosis by modality. Elucidation of ER pathway is helpful to comprehend the pathology of diseases associated with ER stress, and may offer a new approach to the therapy of cancer and neurodegenerative diseases.

• Clinical and Experimental Reproductive Medicine
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• 제49권2호
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• pp.93-100
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• 2022

Objective: The impact of imatinib, a tyrosine kinase inhibitor, on ovarian follicles and several proteins related to follicular function and apoptosis was investigated in mice. Methods: Saline, cyclophosphamide (Cp; 50 or 75 mg/kg), or imatinib (7.5 or 15 mg/kg) was injected once intraperitoneally into female B6D2F1 mice (18 mice in each group). In multiple ovarian sections, the number of various types of follicles and the proportion of good-quality (G1) follicles were counted. The levels of six proteins (anti-Müllerian hormone [AMH], BCL-xL, BAX, acid sphingomyelinase [A-SMase], caspase-3, and α-smooth muscle actin [α-SMA]) within the whole ovaries were quantified using Western blots. Results: Compared to the saline group, a significant reduction of the primordial follicle count was observed in the group treated with imatinib 7.5 and 15 mg/kg, as well as in the group treated with Cp 75 mg/kg. Administration of Cp significantly decreased the proportion of G1 primordial follicles, but administration of imatinib did not. No differences in the AMH, anti-apoptotic BCLX-L, pro-apoptotic BAX, and A-SMase levels in the ovarian tissues were observed among the five groups. However, caspase-3 and α-SMA levels were significantly higher in the imatinib and Cp groups than in the saline group. Conclusion: The administration of imatinib to mice significantly reduced the primordial follicle count and increased the protein levels of caspase-3 and α-SMA. Our findings suggest that imatinib potentially exerts ovarian toxicity via apoptotic processes, similarly to Cp.

• 생명과학회지
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• 제17권8호통권88호
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• pp.1121-1128
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• 2007

초피나무(Z. piperitum) 열매는 다양한 생리활성을 갖고 있으며, 과피 용매추출물은 세균에 대한 강한 항균활성을 갖고 있어, 간장과 된장제조에 초피를 첨가하여 생리활성효과를 조사하였다. 초피를 간장제조에 1%, 2%, 4% 첨가하여 제조하고, 간장추출물을 식중독균(Sta. aureus, Sal. typhimurium, V. parahemolyticus, E. coli 0157:H7)에 대한 항균활성을 조사했다. 초피를 2% 첨가한 간장은 E. coli 0157:H7과 V. parahemolyticuS 의 생육을 각각 70%와 50% 억제했으며, Sal. typhimurium 과 Sta. aureus 에 대한 항균효과는 농도와 간장제조시기에 따라 40% ${\sim}$ 60% 의 생육을 억제했다. 초피된장의 용매추출물에서는 약한 항균활성을 확인할 수 있었다. 초피간장의 추출물을 백혈병세포 U-937에 처리하여 caspase-3 활성으로 항암활성을 조사했다. 그 결과 초피의 량을 1%, 2%, 4% 첨가한 간장의 추출물은 초피를 침가하지 않은 간장보다 caspase-3 활성이 각각 4배, 12배, 15배 활성이 증가하는 것이 확인되어졌다. 이는 초피간장의 추출물이 apoptosis를 유도하는 것을 의미하며, 된장의 용매추출물은 caspase-3 활성에 아무런 영향이 없었다. 초피간장 추출물을 처리한 세포의 죽음이 necrosis인지 apoptosis인지 알아보기 위하여 세포내의 DNA fragmentation을 확인한 결과, 초피간장 추출물을 처리한 세포는 DNA 단편화가 형성되어졌다. 초피틀 1% 첨가한 것에서는 fragmentation을 확인하기 힘들었고, 2%와 4%초피를 첨가한 간장추출물은 DNA fragmentation을 강하게 일으켰다. 초피를 첨가한 간장추출물이 caspase를 활성화시키고, DNA fragmentation을 일으키는 apoptosis 기작에 의해 암세포를 죽음으로 유도하는 초피간장의 항암효과를 입증했다.

• 대한본초학회지
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• 제22권2호
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• pp.35-43
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• 2007

Objectives : The purpose of this study was to investigate the effect of Bo-du-san (BOS) on apoptosis in human gastric cancer cells, SNU-l cells. BOS, a drug preparation consisting of two herbs, that is, Crotonis Fructus (Strychni ignatii Semen, bodu in Korean) and Glycyrrhizae Radix (Glycyrrhizae uralensis FISCH, Gamcho in Korean). Methodss : In this study, methanol extract of BOS was examined for cytotoxic activity on human gastric cancer cells, SNU-1 cells, using XTT assay, with an IC50 value was 0.7 mg/ml and 0.3 mg/ml at 24 hrs and 48 hrs, respectively. Apoptosis induction by BDS in SNU-l cells was verified by the induction of DNA fragmentation, cleavage of poly ADP-ribose polymerase (PARP), and activation of caspase-3, -8 and -9. Inhibitors of caspase-3, -8 and -9 (Ac-DEVD-CHO, Z-IETD-FMK and Z-LEHD-FMK) efficiently blocked BOS-induced cell death of SNU-l. Resultss : BOS-induced cell death was via caspase dependent apoptosis. Moreover, treatment of BOS result in the decrease the G1/S cycle regulation proteins (cyclin D1 and E) expression and increase CDK inhibitor proteins (p21 and p27) expression, and increase apoptotic protein, p53 expression. Thus, BOS induces apoptosis in SNU-1 cells via cell cycle arrested in G1 phase. Conclusions : These results indicated that BOS has some potential for use as an anti-cancer agent.

• Molecules and Cells
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• 제28권6호
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• pp.559-563
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• 2009

Glyceraldehyde-3-phosphate is a key intermediate in several central metabolic pathways of all organisms. Aldolase and glyceraldehyde-3-phosphate dehydrogenase are involved in the production or elimination of glyceraldehyde-3-phosphate during glycolysis or gluconeogenesis, and are differentially expressed under various physiological conditions, including cancer, hypoxia, and apoptosis. In this study, we examine the effects of glyceraldehyde-3-phosphate on cell survival and apoptosis. Overexpression of aldolase protected cells against apoptosis, and addition of glyceraldehyde-3-phosphate to cells delayed apoptosis. Additionally, delayed apoptotic phenomena were observed when glyceraldehyde-3-phosphate was added to a cell-free system, in which artificial apoptotic process was induced by adding dATP and cytochrome c. Surprisingly, glyceraldehyde-3-phosphate directly suppressed caspase-3 activity in a reversible noncompetitive mode, preventing caspase-dependent proteolysis. Based on these results, we suggest that glyceraldehyde-3-phosphate, a key molecule in several central metabolic pathways, functions as a molecule switch between cell survival and apoptosis.

• Parasites, Hosts and Diseases
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• 제52권4호
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• pp.355-365
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• 2014

The enteric protozoan parasite Entamoeba histolytica is the causative agent of human amebiasis. During infection, adherence of E. histolytica through Gal/GalNAc lectin on the surface of the amoeba can induce caspase-3-dependent or -independent host cell death. Phosphorylinositol 3-kinase (PI3K) and protein kinase C (PKC) in E. histolytica play an important function in the adhesion, killing, or phagocytosis of target cells. In this study, we examined the role of amoebic PI3K and PKC in amoeba-induced apoptotic cell death in Jurkat T cells. When Jurkat T cells were incubated with E. histolytica trophozoites, phosphatidylserine (PS) externalization and DNA fragmentation in Jurkat cells were markedly increased compared to those of cells incubated with medium alone. However, when amoebae were pretreated with a PI3K inhibitor, wortmannin before being incubated with E. histolytica, E. histolytica-induced PS externalization and DNA fragmentation in Jurkat cells were significantly reduced compared to results for amoebae pretreated with DMSO. In addition, pretreatment of amoebae with a PKC inhibitor, staurosporine strongly inhibited Jurkat T cell death. However, E. histolytica-induced cleavage of caspase-3, -6, and -7 were not inhibited by pretreatment of amoebae with wortmannin or staurosporin. In addition, we found that amoebic PI3K and PKC have an important role on amoeba adhesion to host compartment. These results suggest that amebic PI3K and PKC activation may play an important role in caspase-independent cell death in Entamoeba-induced apoptosis.

• 한국식품영양과학회지
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• 제45권12호
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• pp.1708-1716
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• 2016

Akt 및 mTOR는 세포 생존에 필수적인 경로로 세포 성장과 증식 등에서 중요한 역할을 하는 것으로 알려져 있다. 본 연구에서는 항암 및 항균 효과가 있는 것으로 알려진 개똥쑥(Artemisia annua L.)에 의한 HepG2 간암세포의 apoptosis 유도 효과를 확인하였다. 본 연구 결과에 의하면 개똥쑥 추출물의 처리 농도가 증가함에 따라 HepG2 세포의 생존율은 억제되었으며, 이는 apoptosis 유도 효과에 의한 것임을 세포의 형태적 변화와 flow cytometry를 통해 확인하였다. 그리고 mitopotential assay와 caspase-3/7 activity assay, western blotting으로 Bcl-2 family 단백질을 확인함으로써 apoptosis 경로 중 내인성 경로(intrinsic pathway)에 의해 apoptosis가 일어남을 알 수 있었다. 이러한 효과는 Akt/mTOR의 활성 저해와 연관이 있었으며 Akt/mTOR의 저해제인 LY294002/rapamycin을 개똥쑥 추출물과 병행처리하였을 경우 개똥쑥 추출물에 의한 apoptosis 효과를 더욱 증대시켰다. 따라서 Akt/mTOR의 저해는 개똥쑥 추출물의 apoptosis 효과를 상승시켰으며 이에 따라 미토콘드리아의 기능 손상과 caspase 활성의 증가를 통해 이루어짐을 확인하였다.