• Archives of Pharmacal Research
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• v.26 no.5
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• pp.405-410
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• 2003

Vitamin K-related analogs induce growth inhibition in various cancer cell lines. A naphthoquinone analog, termed 2,3-dichloro-5, 8-dihydroxy-1,4-naphthoquinone (DDN), induces apoptosis in human promyeloid leukemic HL-60 cells, and shows antitumor activity in vivo. Following treatment with DDN, evidence of apoptosis, including DNA fragmentation and cleavage of poly ADP ribose polymerase (PARP), was observed. DDN induced an upregulation of proapoptotic Bax protein, and Bid cleavage. Antiapoptotic Bcl-2 protein levels were not changed by DDN, but the expression of Bcl-xL was decreased. In addition, DDN reduced the mass of solid tumor in the Sarcoma 180 tumor-bearing mouse model. These results indicate that DDN exerts antitumor activity, which appears to be related to the induction of apoptosis by regulating Bcl-2 family proteins.

• The Journal of Internal Korean Medicine
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• v.35 no.1
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• pp.79-91
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• 2014

Objectives : To investigate the anti-cancer effect of Palbohoichoon-tang (PBHCT) extracts. Methods : The cell viability was assessed by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide (MMT) assay and cell morphological changes were microscopically analyzed after staining with $10{\mu}M$ 2-[4-amidinophenyl]-6-indolecarbamidine dihydrochloride (DAPI) and TUNEL. We also analyzed expression of Bcl2, $Bcl_{xL}$, Bax, procaspase-3, procaspase-9, and procyclic acidic repetitive protein (PARP) by western blot method. Results : Observations showed that PBHCT induced the apoptotic cell death proved by increased sub-G1 phase cell population, apoptotic body formation and chromatin condensation. Western blot analysis of total cell lysates revealed that the PBHCT induced cleavage of caspase-9, caspase-3 and poly (ADP-ribose) polymerase (PARP). In addition, PBHCT dose-dependently increased the activity of caspase-9, caspase-3 and PARP-1. Furthermore, PBHCT reduced anti-apoptotic Bcl2, $Bcl_{xL}$ expression which contributed to the loss of mitochondrial membrane potential and the activations of caspase-9 and caspase-3. Conclusions : These findings suggest that PBHCT exerts anti-cancer effects on human neuroblastoma SH-SY5Y cells by inducing apoptotic death via down-regulation of anti-apoptotic proteins such as Bcl2 and $Bcl_{xL}$, up-regulation of pro-apoptotic proteins such as Bax, and activation of caspase cascades and PARP-1.

• Proceedings of the KSAR Conference
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• 2001.03a
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• pp.52-52
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• 2001

Many nucleoside diphosphate (NDP) kinases are ubiquitous enzymes responsible for the exchange of ${\gamma}$-phosphates between tri- and diphosphonucleosides. The catalytic Many nucleoside diphosphate (NDP) kinases are ubiquitous enzymes responsible for the exchange of ${\gamma}$-phosphates between tri- and diphosphonucleosides. The catalytic reaction follows a ping-pong mechanism in which the enzyme is transiently phosphorylated on a histidine residue conserved in all nucleoside diphosphate kinases. Beside their role in nucleotide synthesis, these enzymes present additional functions, possibly independent of catalysis, in processes such as differentiation, cell growth, tumor progression, metastasis and development. To clone murine nm23-M5, several expressed sequence tags (ESTs) of the GenBank data base, selected according to their homology to nm23-H5 cDNA, reconstituted a complete open reading frame (GenBank AF222750). To test whether murine NDPKs (1, 2, 3, 4, 5, and 6) can inhibit Bax-mediated toxicity in yeast, co-transformation was performed respectively. The yeast S.cerevisiae was transformed with a copy expression plasmid containing the histidine selection marker and expressing murine Bax under the control of a galactose-inducible promoter. Several clones were selected and found to be growth inhibited when Bax expression was induced with galactose. A representative clone was transformed again with a copy expression plasmid containing the tryptophane selection marker and expressing either murine Bcl-xL or NDPK under the control of a galactose-inducible promoter. Several subclones of the double-transformants were selected and characterized. The ability of Bcl-xL and NDPKs to suppress Bax-mediated toxicity was determined by growing yeast cells overnight in galactose media and spot-testing on galactose plates starting with an equal number of yeast cells as determined by taking the OD$_{600}$. Ten-fold serial dilutions were used in the spot-test. Plates were grown at 3$0^{\circ}C$ for 2-3 days. All murine NDPKs suppressed Bax dependent apoptosis. Futher study will be peformed whether Bax-toxicity inhibition was caused by NDP kinase activity or additional function.n.

• Clinical and Experimental Pediatrics
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• v.46 no.10
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• pp.989-995
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• 2003

Purpose : Dexamethasone is frequently administered to prevent or treat chronic lung disease in human neonates who are also prone to hypoxic-ischemic(HI) insults. Recently, meta-analysis of the follow-up studies reveals a significantly increased odd ratio for the occurrence of cerebral palsy or an abnormal neurologic outcome, and there is conflicting evidence regarding the impact of dexamethasone exposure on HI brain injury. This study was conducted to explore the effect of post-HI dexamethasone administration on neuronal injury in neonatal rats. Methods : HI was produced in seven-day-old rats by right carotid artery ligation followed by two hours of 8% oxygen exposure. At the end of HI, the animals were injected intraperitoneally either with dexamethasone(0.5 mg/kg) or saline. Neuronal injury was assessed seven days after the HI by the area of infarction, TUNEL reactivity, Bcl-2 and Bax expression in brain. Results : Post-insult dexamethasone administration resulted in reduction of weight gain and a higher mortality rate during seven days after HI. Dexamethasone treatment revealed no effect on the size of brain infarction induced by HI. Bax protein expression increased in dexamethasone treated brain but Bcl-2 protein expression and TUNEL reactivity revealed no significant differences between dexamethasone treated and non treated brain. Increased Bax protein expression suggest upregulation of the apoptosis by dexamethasone. Conclusion : The result suggests the adverse role of Post-HI administration of dexamethasone in neonatal HI.

• Proceedings of the Korean Society of Developmental Biology Conference
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• 2003.10a
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• pp.114-114
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• 2003

Epidermal growth factor (EGF) induces well-documented mitogenic and differentiating effects on murine and bovine preimplantation embryos. However, the effects of EGF on apoptosis and implantation-related gene expression in bovine embryos developing in vitro have not been evaluated. The objective of this study was to determine the effects of exogenous EGF in the presence and absence of BSA on the preimplantation development of bovine embryos. In addition, we measured cell number, apoptosis, and expression of apoptosis and implantation-related genes of the blastocysts that developed in these culture conditions. In vitro produced bovine embryos were randomly cultured in the same medium containing 0 or 10 ng/ml EGF in the presence and absence of 0.8% BSA. More 2-cell embryos developed into blastocysts at day 7 when BSA was present than when BSA was absent. The addition of 10 ng/$m\ell$ EGF into the medium did not significantly increase the developmental rate and the cell numbers per blastocyst. However, addition of EGF in the presence of 0.8% BSA significantly reduced the degree of apoptosis in the blastocysts (P<0.01). To investigate whether EGF modulates mRNA expression of apoptosis-related genes, mRNA was prepared from single blastocysts and each preparation was subjected to RT-PCR for Bcl-2 and Bax transcripts. EGF did not alter the relative abundance of Bax gene expression in the presence of BSA, but increase Bcl-2 (P<0.01) The relative abundance of Interferon tau expression was increased by EGF treatment in the presence of BSA. These results suggest that EGF and BSA synergistically enhance Bcl-2 and interferone tau gene expression, which may result in a net increase in viability in bovine embryos.

• Proceedings of the Korean Society of Developmental Biology Conference
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• 2003.10a
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• pp.99-99
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• 2003

Epidermal growth factor (EGF) induces well-documented mitegenic and differentiating effects on murine and bovine preimplantation embryos. However, the effects of EGF on apoptosis and implantation-related gene expression in bovine embryos developing in vitro have not been evaluated. The objective of this study was to determine the effects of exogenous EGF in the presence and absence of BSA on the preimplantation development of bovine embryos. In addition, we measured cell number, apoptosis, and expression of apoptosis and implantation-related genes of the blastocysts that developed in these culture conditions. In vitro produced bovine embryos were randomly cultured in the same medium containing 0 or 10 ng/$m\ell$ EGF in the presence and absence of 0.8% BSA. More 2-cell embryos developed into blastocysts at day 7 when BSA was present than when BSA was absent. The addition of 10 ng/$m\ell$ EGF into the medium did not significantly increase the developmental rate and the cell numbers per blastocyst. However, addition of EGF in the presence of 0.8% BSA significantly reduced the degree of apoptosis in the blastocysts (P< 0.01). To investigate whether EGF modulates mRNA expression of apoptosis-related genes, mRNA was prepared from single blastocysts and each preparation was subjected to RT-PCR for Bcl-2 and Bax transcripts. EGF did not alter the relative abundance of Bax gene expression in the presence of BSA, but increase Bcl-2 (P < 0.01). The relative abundance of Interferon tau expression was increased by EGF treatment in the presence of BSA. These results suggest that EGF and BSA synergistically enhance Bcl-2 and interferone tau gene expression, which may result in a net increase in viability in bovine embryos.

• Biomolecules & Therapeutics
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• v.12 no.2
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• pp.85-91
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• 2004

In all attempt to elucidate the role of apoptosis in drug resistance, cisplatin-resistant human chronic myelogenous leukemia (CML) K562 cells (K562/CDDP) were established and compared with drug sensitive parent cells (K562) in the induction of apoptosis. K562/CDDP cells were 5-fold more resistant to cisplatin compared to K562 cells. In addition, K562/CDDP cells were significantly more resistant to apoptois as judged by DNA fragmentation and DAPI staining. K562/CDDP cells exhibited decreased proleolytic activity of caspase-3 and this was further demonstrated by decreased cleavage of its substrate poly (ADP-ribose) polymerase (PARR- Western blot analysis showed that K562/CDDP cells had longer sustained levels of BCL-$X_L$ whereas no difference was noted in the level of Bcl-2. the translocation of Bax to mitochondria was significantly delayed in K562/CDDP cells. These results suggest that the reduced translocation of Bax and the sustained expression of BCL-$X_L$ may cause resistance to apoptosis through prevention of mitochondria release of cytochrome c, which subsequently induces reduction of caspase-3 activity and that this response is partly responsible for the acquired resistance to cisplatin ill K562 cells.

• Toxicological Research
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• v.19 no.1
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• pp.67-72
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• 2003

Cholestatic liver injury results from the accumulation of toxic bile salts within the liver. The aim of the present study is to elucidate the changes in expression and cellular localization of apoptosis related proteins in the liver of bile duct-ligated (BDL) rat. Extrahepatic cholestasis was induced by double ligation of the common bile duct and cut between the ligatures. Animals were sacrificed at day 3 and at week 1, 2, 4, 6, and 8 after BDL. The number of TUNEL positive cells was increased significantly after 3 days of BDL, decreased over 2 weeks and remained constant thereafter. Fas expression was not changed and activation of caspase 8 did not occur. Fas immunoreactivity was exclusively observed in the cytoplasm of hepatocytes, indicating that Fas expressed in rat hepatocytes is a soluble form. Hepatocyte apoptosis was associated with Bax expression, which showed a peak at day 3 and decreased over time gradually. Immnunostaining of Bax was observed in hepatocytes and bile duct epithelial cells (BEC) of control and BDL rats. Bcl-2 was increased over time in BDL rats. These results suggest that apoptosis of hepatocytes in BDL rats is independent of Fas and controlled by Bax expression.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.11
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• pp.5753-5757
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• 2012

Apoptosis, also known as cell suicide or programmed cell death, removes unwanted and genetically damaged cells from the body. Evasion of apoptosis is one of the major characteristic features of rapidly proliferating tumor cells. Chemopreventive agents inhibit or suppress tumor formation through apoptotic induction in target tissues. The aim of the present study was to investigate the pro-apoptotic potential of lupeol during 7,12-dimethylbenz(a) anthracene (DMBA) induced hamster buccal pouch carcinogenesis. Topical application of 0.5% DMBA three times a week for 14 weeks in the buccal pouches of golden Syrian hamsters resulted in oral squamous cell carcinoma. The expression pattern of apoptotic markers was analyzed using immunohistochemistry (p53, Bcl-2, Bax) and ELISA reader (caspase 3 and 9). In the present study, 100% tumor formation with defects in apoptotic markerexpression pattern was noticed in hamsters treated with DMBA alone. Oral administration of lupeol at a dose of 50mg/kg bw completely prevented the formation oral tumors as well as decreased the expression p53 and Bcl-2, while increasing the expression of Bax and the activities of caspase 3 and 9. The present study thus indicated that lupeol might inhibit DMBA-induced oral tumor formation through its pro-apoptotic potential in golden Syrian hamsters.

• The Journal of Internal Korean Medicine
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• v.33 no.4
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• pp.465-475
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• 2012

Objectives : This study was performed to investigate the antineoplastic effect of ethanol extracts from Sanguisorbae Radix on cholangiocarcinoma cells that was established from biliary tract cancer tissue. Materials and Methods : Two cholangiocarcinoma cell lines, SNU-1079 and SNU-1196, were studied. The mRNA expression of Caspase 3, 8, 9, Bcl-2, Bax, P53, and P21 was examined by RT-PCR. Cell viability was determined by MTT assay. The cell cycle was analyzed by flow cytometry and apoptosis by cell death detection ELISA kit. Results : Proliferation of SNU-1079 and SNU-1196 was inhibited by Sanguisorbae Radix treatment in a dose-dependent manner. All cells treated with Sanguisorbae Radix showed increased dose- and time-dependent apoptosis. The expression of caspase 3, 8, 9, p53, and p21 was increased in all cells after the treatment of Sanguisorbae Radix. The expression of Bcl-2 was decreased in SNU-1196 and Bax expression was increased in all cells after the treatment of Sanguisorbae Radix. Conclusions : These results suggest that Sanguisorbae Radix would be beneficial in the treatment of cholangiocarcinoma.