• The Journal of Internal Korean Medicine
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• v.21 no.1
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• pp.65-73
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• 2000

Objectives : The effects of cotreatment of adriamycin and ethanol extract of herb (Palgin-tang hab Hwajuck-hwan a traditional medicine for cancer treatment in oriental medicine) on the induction of apoptotic cell death were investigated in human liver origin cell lines, Chang. Methods : Chang(ATCC) liver cells were cultured in RPMI-1640(Gibco SRL Co, Gaithersburg, MD) badge including 10% fetal bovine serum. Chang liver cells were treated with various concentrations(from 10 to $0.16{\mu}l$) of adriamycin and herb extract(from 500 to $31.25{\mu}l$) After 48h later, the cells were tested for viability by Crystal violet staining assay. Adriamycin and Herb extract induced ladder pattern of DNA fragmentation in Chang cells. Genomic DNA was isolated and separated on 1.5% agarose gels. The DNA was stained with ethidium bromide and visualized under UV light. Results : The death of Chang cells was synergistically induced by the cotreatment of adriamycin and ethanol extract of herb. In addition, the cotreatment-induced cell death of Chang cells was mediated by apoptotic death signal processes. The phosphotransferase activity of JNK1 remained in a basal level in Chang cells which was treated individually with the adriamycin and ethanol extract of herb. However, it was markedly increased in Chang cells which was cotreated with adriamycin and ethanol extract of herb. In addition, the expression of Fas and FasL was markedly induced by the cotreatment of adriamycin and herb extract. For a while, the expression of Sax was a eminently increased by the ethanol extract of herb. However, Scl2 expression was not affected by the individual or cotreatment of adriamycin and herb extract. Conclusions : our results suggest that the cotreatment of adriamycin aM ethanol extract of herb induces synergistic apoptotis of human liver origin Chang cells via the upregulation of JNK, Fas, FasL and Bax.

• Herbal Formula Science
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• v.15 no.1
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• pp.213-228
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• 2007

Objectives : The purpose of this report was to investigate the chemotherapeutic effect of Bujeonghangamtang against cancer cells. Materials and Methods : Various cancer cell lines including PANC-1, C6 glioma, SH-SY5Y, HepG2, and MCF-7 cells, were used. Apoptosis was determined by DAPI nuclei staining and flow cytometry in PANC-1 cells treated with 1 mg/ml Bujeonghangamtang for 48 hr. Expression of cell cycle arrest mediators including, cdc2p34 and cyclin B1 proteins were measured by Western blot analysis. Mitochondrial membrane potential was measured by fluorescence staining with JC-1, rhodamine 123. Result : Bujeonghangamtang induced the apoptosis of PANC-1, which was characterized as nucleic acid and genomic DNA fragmentation, chromatin condensation, and sub-G0/G1 fraction of cell cycle increase. but not C6 glioma, SH-SY5Y, HepG2, and MCF-7 cells. PANC-1 cells were markedly sensitive to Bujeonghangamtang. Treatment with Bujeonghangamtang resulted in the decreased expression of cdc2p34 and cyclin B1. Treatment with Bujeonghangamtang also increased the ROS production and induced mitochondrial dysfunction. Conclusion : Bujeonghangamtang exerted cytotoxicity against human Pancreatic cancer cells via cell cycle arrest-mediated apoptotic signaling including ROS production and mitochondrial dysfunction. Our data suggest that Bujeonghangamtang may be an important modulator of chemosensitivity of cancer cells against anticancer chemotherapeutic agents.

• Journal of Physiology & Pathology in Korean Medicine
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• v.28 no.3
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• pp.310-316
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• 2014

Oldenlandia diffusa, Cremastra appendiculata and Fritillaria thunbergii are widely distributed in the Korea, China and Japan, and has been used in traditional medicine for various diseases, such as pharyngolaryngitis, tonsillitis, goiter and stomach ulcer. However, the anti-cancer activities in human breast cancer have not been clearly elucidated yet. In this study, it was compared the in vitro cytotoxic effects of single and complex treatment of O. diffusa, C. appendiculata and F. thunbergii. We treat human breast cancer MCF-7 cells with O. diffusa, C. appendiculata and F. thunbergii. And we evaluated viability, growth inhibition, morphological changes, apoptotic bodies formation, measurement of the cell cycle and formation of DNA fragmentation of these cells. It was found that single treatment of O. diffusa could inhibit the cell proliferation in human breast cancer MCF-7 cells. However, complex treatment of O. diffusa, C. appendiculata and F. thunbergii is weakly or not affect the cell proliferation of MCF-7 cells. And anti-proliferative effects of O. diffusa in MCF-7 cells was associated with G1 arrest of cell cycle. These findings suggest that O. diffusa may be a potential chemotherapeutic agent for the control of human breast cancer cells and further studies will be needed to identify the molecular mechanisms.

• Advances in Traditional Medicine
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• v.10 no.4
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• pp.254-262
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• 2010

Artemisia capillaris (A. capillaries) is known to play roles in many cellular events, such as cell proliferation, differentiation, and apoptosis. We investigated the antifibrogenic efficacy of A. capillaris in the immortalized human hepatic stellate cell line LX2. Cell proliferation was determined by the MTT assay. Cell cycle was analyzed by the flow cytometry. Apoptotic cells were measured using a cell death detection ELISA. Caspase activity was detected by a colorimetric assay. The mRNA level of Bcl-2 and Bax mRNA were measured by real-time PCR. MEK and ERK protein were detected by Western blot analysis. We provide evidence that A. capillaris induces cell cycle arrest, apoptosis, and potently inhibits the mitogen-activated protein kinase pathway. A. capillaris inhibited cell proliferation of LX2 cells in a dose- and time-dependent manner, increased the apoptosis fraction at cell cycle analysis with an accompanying DNA fragmentation, and resulted in a significant decrease in Bcl-2 mRNA levels and an increase in Bax expression. Exposure of LX2 cells to A. capillaris induced caspase-3 activation, but co-treatment of A. capillaris with the pan-caspase inhibitor Z-VAD-FMK, and the caspase-3 inhibitor Z-DEVE-FMK, blocked apoptosis. A. capillaris down-regulated Mcl-1 protein levels and inhibited phosphorylation of MEK/ERK, suggesting that it mediates cell death in LX2 cells through the down-regulation of Mcl-1 protein via a MEK/ERK-independent pathway.

• The Journal of Korean Medicine
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• v.19 no.2
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• pp.337-372
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• 1998

To evaluate the effects of Injinchunggantang-derivative on cell viability, cell cycle progression, and apoptosis, MTT assay, cell cycle analysis, Cpp32 protease assay, DNA fragnemtation assay, quantitative RT-PCR, and Western blotting were performed. The results were as followes. In MTT assay, etoposide+Injinchunggantang-derivative-treated cells as well as Injinchunggantang-derivative-treated cells showed higher viability than etoposide-treated cells with no time-concentration-dependence, which implied that Injinchunggantang-derivative has hepato-protective effect Cell cycle analysis showed that Injinchunggantang-derivative has no significant effect on the cell cycle. Cpp32 protease assav and DNA fragmentation assay Injinchunggantang-derivative carry inhibitory effects on apoptosis induction. It was suggested that Injinchunggantang-delivative might regulate the cell cycle, in particular $G_1$ checkpoint by blocking p53 and Watl pathway. Injinchunggantang-derivative inhibited the mRNA expressions of Cpp32, Fas, and Bcl-2, which could result in inhibition of apoptosis. These results imply that Injinchunggantang-derivative increases hepatocyte viability, and protects hepatocyte from damage by regulating the expression of genes associated with cell cycle and apoptosis, which explains the mechanism of the clinical effect of Injinchunggantang-derivative on liver diseases.

• Journal of Physiology & Pathology in Korean Medicine
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• v.20 no.6
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• pp.1459-1466
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• 2006

The water extract of Hwansodan has been traditionally used for treatment of ischemic brain damage in oriental medicine. However, little is known about the mechanism by which the water extract of Hwansodan rescues cells from neurodegenerative disease. PC12 pheochromocytoma cells have been used extensively as a model for studying the cellular and molecular mechanisms of neuronal cell damages. Under deprivation of growth factor and ischemic injury, PC12 cells spontaneously undergoes apoptotic cell death. Serum and glucose deprivation markedly decreased the viability of PC12 cells, which was characterized with apparent apoptotic features such as membrane blebbing as well as fragmentation of genomic DNA and nuclei. However, the aqueous extract of Hwansodan significantly reduced serum and glucose deprivation-induced cell death and apoptotic characteristics through reduction of intracellular peroxide generation. Pretreatment of Hwansodan also ingibited the activation of caspase-3, in turn, degradation of ICAD/DFF45 was completely abolished in serum and glucose deprivated cells. Furthermore, pretreatment of Hwansodan obviously increased heme oxygenase 1 (HO-1) expression in PC12 cells. Taken together, the data suggest that the protective effects of Hwansodan against serum and glucose deprivation induced oxidative injuries may be achieved through the scavenging of reactive oxygene species accompanying with HO-1 induction.

• Journal of Physiology & Pathology in Korean Medicine
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• v.20 no.6
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• pp.1516-1523
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• 2006

The water extract of Danchunhwan(DCH) has been traditionally used for treatment of dementia damage in oriental medicine. However, little is known about the mechanism by which the water extract of DCH rescues cells from neurodegenerative disease such as Alzheimer's disease. This study was designed to investigate the protective mechanisms of DCH on ${\beta}$-amyloid or $H_2O_2$-induced cytotoxicity in SH-SY5Y neuronblastoma cells. ${\beta}$-amyloid and $H_2O_2$ markedly decreased the viability of SH-SY5Y cells, which was characterized with apparent apoptotic features such as membrane blebbing as well as fragmentation of genomic DNA and nuclei. However, the water extract of DCH significantly reduced both ${\beta}$-amyloid or $H_2O_2$-induced cell death and apoptotic characteristics through reduction of intracellular peroxide generation. Also, the water extract of DCH prevented prevented the mitochondrial dysfunction including the disruption of mitochondria membrane permeability transition (MPT) and the perturbation in Bcl-2 family protein expressions in $H_2O_2$-treated SH-SY5Y cells.

• The Journal of Internal Korean Medicine
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• v.21 no.2
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• pp.259-266
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• 2000

Objectives : The effects of aqueous extracts of Cortex ulmi pumilae (a traditional medicine for cancer treatment in oriental medicine) on the induction of apoptotic cell death were investigated in human liver origm hepatoma cell lines, HepG2. Methods : The death of HepG2 cells was markedly induced by the addition of extracts of Cortex ulmi pumilae in a dose-dependent manner. The apoptotic characteristic ladder pattern of DNA strand break was not observed in cell death of HepG2. In addition, it was not shown nucleus chromatin condensation and fragmentation under hoechst staining. However, by the using annexin V staining assay, externalizations of phosphatidylserine in HepG2 cell which were treated with Cortex ulmi pumilae extracts were detected in the early time (at 9 hr after extract treatment). Furthermore, LDH release was not detected in this early stage. Therefore, Cortex ulmi pumilae extracts-induced cell death of HepG2 cells is mediated by apoptotic death signal processes. Result : The activity of caspase 3-like proteases remained in a basal level in HepG2 cells which treated with the extract of Cordyceps sinensis. However, it was markedly increased in HepG2 cells which treated with two extracts of Cortex ulmi pumilae (C.U.P.-C, C.U.P.-K) which were differently extracted (respectively, 2.3 and 3.3 fold). On a while, the phosphotransferase activities of JNK1 was markedly induced in HepG2 cells which were treated with two extracts of Cortex ulmi pumilae. On the contrary, the activation of transcriptional activator, activating protein1(AP-1) and NF-kB were severely decreased by these two extracts of Cortex ulmi pumilae (C.U.P.-C, C.U.P.-K). In addition, antioxidants (GSH and NAC) and intracellular $Ca2^+$ level regulator (Bapta/AM and Thapsigargin) did not affect Cortex ulmi pumilae extracts-induced apoptotic death of HepG2 cells. Conclusions : In conclusion, our results suggest that two extracts of Cortex ulmi pumilae (C.U.P.-C, C.U.P.-K) induces the apoptotic death of human liver origin hepatoma HepG2 cells via activation of caspase 3-like proteases as well as JNK1, and inhibition of transcriptional activators, AP-1 and $NK-{\kappa}B$.

• The Journal of Internal Korean Medicine
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• v.29 no.1
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• pp.130-148
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• 2008

Objective : This study was designed to investigate the effect of the water extract of Gamisamgibopae-tang(GMSGBPT), an oriental herbal formulation, on the growth of NCI-H460 and A549 human non-small-cell lung cancer cell lines. Methods : Cytotoxicity and cell morphology were evaluated by MTT assay and inverted microscope, respectively. Apoptosis was detected using agarose gel electrophoresis and flow cytometer. The expression levels of mRNAs and proteins of target genes were determined by RT-PCR and western blot analyses, respectively Result and Conclusion : We found that exposure of A549 cells to GMSGBPT resulted in the growth inhibition in a dose-dependent manner as measured by MTT assay, but GMSGBPTdid not affect the growth of NCI-H460 cells. The anti-proliferative effect of GMSGBPT treatment in A549 cells was associated with morphological changes, formation of apoptotic bodies and DNA fragmentation, and flow cytometry analysis confirmed that GMSGBPT treatment increased the populations of apoptotic-sub G1 phase. Growth inhibition and apoptotic cell death by GMSGBPT were connected with a up-regulation of cyclin-dependent kinase inhibitor p21 (WAF1/CIP1) mRNA and protein in a tumor suppressor p53-independent fashion. However GMSGBPT treatment did not affect other growth regulation-related genes such as early growth response-1 (Egr-1), nonsteroidal anti-inflammatory drug (NSAID)-activated gene-1 (NAG-1), inducible nitric oxide synthase (iNOS), cyclooxygenases (COXs), telomere-regulatory factors in A549 orNCI-H460 cells. Taken together, these findings partially provide novel insights into the possible molecular mechanism of the anti-cancer activity of GMSGBPT.

• Journal of Physiology & Pathology in Korean Medicine
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• v.17 no.6
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• pp.1383-1392
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• 2003

Apoptosis is a morphologically and biochemically district form of cell death that occurs in many different cell types in a wide variety of organisms. Albizzia julibrissin belonging the family Leguminosae has been used for the treatment of contusion, sore throat, amnesia, and insomnia in oriental traditional medicine. This study investigates whether the water extract of A. julibrissin induce apoptotic cell death in Jurkat T-acute lymphoblastic leukemia (ALL) cells. Jurkat cells were increased inhibitions of cell viability in a concentration-dependent manner by A. julibrissin. This herbal medicine also caused apoptosis as measured by cell morphology and DNA fragmentation. The capability of A. julibrissin to induce apoptosis was associated with proteolytic cleavage of specific target proteins such as poly (ADP-ribose)polymerase (PARP) and beta-catenin proteins suggesting the possible involvement of caspases. Our result showed that Bcl-2 and Bax protein levels were not changed in all A. julibrissin-treated groups compared to control group. These results suggest that A. julibrissin-mediated apoptosis is independent with Bcl-2 related signaling pathway in this cells. The purpose of the present study is also to investigate the Effect of A. julibrissin on cell cycle progression. Our results showed that G1 checkpoint related gene products (cyclin D1, cyclin dependent kinase 4, retinoblastoma, E2F1) were decreased in their protein levels in a dose-dependent manners after treatment of the extract. These results indicate that the increase of apoptotic cell death by A. julibrissin may be due to the inhibition of cell cycle progression in wild type p53-lacking Jurkat cells.