• Radiation Oncology Journal
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• v.30 no.2
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• pp.78-87
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• 2012

Purpose: Troglitazone (TRO) is a peroxisome proliferator-activated receptor ${\gamma}$ ($PPAR{\gamma}$) agonist. TRO has antiproliferative activity on many kinds of cancer cells via G1 arrest. TRO also increases $Cu^{2+}/Zn^{2+}$-superoxide dismutase (CuZnSOD) and catalase. Cell cycle, and SOD and catalase may affect on radiation sensitivity. We investigated the effect of TRO on radiation sensitivity in cancer cells in vitro. Materials and Methods: Three human cervix cancer cell lines (HeLa, Me180, and SiHa) were used. The protein expressions of SOD and catalase, and catalase activities were measured at 2-10 ${\mu}M$ of TRO for 24 hours. Cell cycle was evaluated with flow cytometry. Reactive oxygen species (ROS) was measured using 2',7'-dichlorofluorescin diacetate. Cell survival by radiation was measured with clonogenic assay. Results: By 5 ${\mu}M$ TRO for 24 hours, the mRNA, protein expression and activity of catalase were increased in all three cell lines. G0-G1 phase cells were increased in HeLa and Me180 by 5 ${\mu}M$ TRO for 24 hours, but those were not increased in SiHa. By pretreatment with 5 ${\mu}M$ TRO radiation sensitivity was increased in HeLa and Me180, but it was decreased in SiHa. In Me180, with 2 ${\mu}M$ TRO which increased catalase but not increased G0-G1 cells, radiosensitization was not observed. ROS produced by radiation was decreased with TRO. Conclusion: TRO increases radiation sensitivity through G0-G1 arrest or decreases radiation sensitivity through catalase-mediated ROS scavenging according to TRO dose or cell types. The change of radiation sensitivity by combined with TRO is not dependent on the PPAR ${\gamma}$ expression level.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.1
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• pp.489-494
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• 2014

Background: Psychiatric patients appear to be at lower risk of cancer. Some antipsychotic drugs might have inhibitory effects on tumor growth, including penfluridol, a strong agent. To test this, we conducted a study to determine whether penfluridol exerts cytotoxic effects on tumor cells and, if so, to explore its anti-tumor mechanisms. Methods: Growth inhibition of mouse cancer cell lines by penfluridol was determined using the 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. Cytotoxic activity was determined by clonogenic cell survival and trypan blue assays. Animal tumor models of these cancer cells were established and to evaluate penfluridol for its anti-tumor efficacy in vivo. Unesterified cholesterol in cancer cells was examined by filipin staining. Serum total cholesterol and tumor total cholesterol were detected using the cholesterol oxidase/p-aminophenazone (CHOD-PAP) method. Results: Penfluridol inhibited the proliferation of B16 melanoma (B16/F10), LL/2 lung carcinoma (LL/2), CT26 colon carcinoma (CT26) and 4T1 breast cancer (4T1) cells in vitro. In vivo penfluridol was particularly effective at inhibiting LL/2 lung tumor growth, and obviously prolonged the survival time of mice bearing LL/2 lung tumors implanted subcutaneously. Accumulated unesterified cholesterol was found in all of the cancer cells treated with penfluridol, and this effect was most evident in LL/2, 4T1 and CT26 cells. No significant difference in serum cholesterol levels was found between the normal saline-treated mice and the penfluridol-treated mice. However, a dose-dependent decrease of total cholesterol in tumor tissues was observed in penfluridol-treated mice, which was most evident in B16/F10-, LL/2-, and 4T1-tumor-bearing mice. Conclusion: Our results suggested that penfluridol is not only cytotoxic to cancer cells in vitro but can also inhibit tumor growth in vivo. Dysregulation of cholesterol homeostasis by penfluridol may be involved in its anti-tumor mechanisms.

• Asian Pacific Journal of Cancer Prevention
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• v.16 no.7
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• pp.3035-3042
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• 2015

Background: Isorhamnetin (Iso), a novel and essential monomer derived from total flavones of Hippophae rhamnoides that has long been used as a traditional Chinese medicine for angina pectoris and acute myocardial infarction, has also shown a spectrum of antitumor activity. However, little is known about the mechanisms of action Iso on cancer cells. Objectives: To investigate the effects of Iso on A549 lung cancer cells and underlying mechanisms. Materials and Methods: A549 cells were treated with $10{\sim}320{\mu}g/ml$ Iso. Their morphological and cellular characteristics were assessed by light and electronic microscopy. Growth inhibition was analyzed by MTT, clonogenic and growth curve assays. Apoptotic characteristics of cells were determined by flow cytometry (FCM), DNA fragmentation, single cell gel electrophoresis (comet) assay, immunocytochemistry and terminal deoxynucleotidyl transferase nick end labeling (TUNEL). Tumor models were setup by transplanting Lewis lung carcinoma cells into C57BL/6 mice, and the weights and sizes of tumors were measured. Results: Iso markedly inhibited the growth of A549 cells with induction of apoptotic changes. Iso at $20{\mu}g/ml$, could induce A549 cell apoptosis, up-regulate the expression of apoptosis genes Bax, Caspase-3 and P53, and down-regulate the expression of Bcl-2, cyclinD1 and PCNA protein. The tumors in tumor-bearing mice treated with Iso were significantly smaller than in the control group. The results of apoptosis-related genes, PCNA, cyclinD1 and other protein expression levels of transplanted Lewis cells were the same as those of A549 cells in vitro. Conclusions: Iso, a natural single compound isolated from total flavones, has antiproliferative activity against lung cancer in vitro and in vivo. Its mechanisms of action may involve apoptosis of cells induced by down-regulation of oncogenes and up-regulation of apoptotic genes.

• BMB Reports
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• v.51 no.2
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• pp.92-97
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• 2018

B cell leukemia/lymphoma 3 (Bcl3) plays a pivotal role in immune homeostasis, cellular proliferation, and cell survival, as a co-activator or co-repressor of transcription of the $NF-{\kappa}B$ family. Recently, it was reported that Bcl3 positively regulates pluripotency genes, including Oct4, in mouse embryonic stem cells (mESCs). However, the role of Bcl3 in the maintenance of pluripotency and self-renewal activity is not fully established. Here, we report the dynamic regulation of the proliferation, pluripotency, and self-renewal of mESCs by Bcl3 via an influence on Nanog transcriptional activity. Bcl3 expression is predominantly observed in immature mESCs, but significantly decreased during cell differentiation by LIF depletion and in mESC-derived EBs. Importantly, the knockdown of Bcl3 resulted in the loss of self-renewal ability and decreased cell proliferation. Similarly, the ectopic expression of Bcl3 also resulted in a significant reduction of proliferation, and the self-renewal of mESCs was demonstrated by alkaline phosphatase staining and clonogenic single cell-derived colony assay. We further examined that Bcl3-mediated regulation of Nanog transcriptional activity in mESCs, which indicated that Bcl3 acts as a transcriptional repressor of Nanog expression in mESCs. In conclusion, we demonstrated that a sufficient concentration of Bcl3 in mESCs plays a critical role in the maintenance of pluripotency and the self-renewal of mESCs via the regulation of Nanog transcriptional activity.

• The Korea Journal of Herbology
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• v.28 no.4
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• pp.7-16
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• 2013

Objectives : Brassica campestris var narinosa (BCN), Canavalia gladiata DC semen (CGD) and their combinational prescription (BCN+CGD) have been use to demonstrate to regulate hematopoiesis. In the current study, we investigated whether Brassica campestris var narinosa, Canavalia gladiata DC semen and their combinational prescription is related to hemato-potentiating function using Sca-$1^+$ hematopoietic stem cells (Sca-$1^+HSCs$) as a testing system. Methods : Sca-$1^+HSCs$ isolated from femur in C57bl/6 mice with leukopenia and thrombocytopenia induced by cyclophosphamide (CTX). Then, Real-time PCR was performed to measure the mRNA expression, ELISA and haematopoiesis-related gene (EPO, TPO, IL-3, SCF, c-kit, GM-CSF), the phosphorylation of JAK2, GATA-1 and STAT-5a/b were observed by western blot, and the numbers of $CD117^+/Sca-1^+$ cell and the number of granulocyte erythrocyte monocyte macrophage colony-forming units (CFU-GEMM) and erythroid burst forming units (BFU-E), semisolid clonogenic assay was performed. Result : When Sca-$1^+HSCs$ were treated with Brassica campestris var narinosa, Canavalia gladiata DC semen and their combinational prescription with rIL-3/rSCF, the expression of haematopoiesis-related (EPO, TPO, IL-3, SCF, c-kit, and GM-CSF) were significantly increased at the levels of mRNA as well as production in Sca-$1^+HSCs$. Additionally, CGS enhanced phosphorylation of JAK2, GATA-1, and signal transducer and activator of transcription-5a/b (STAT-5a/b) in Sca-$1^+HSCs$. Furthermore, their combinational prescription (BCN+CGD) significantly enhanced the growth rate of granulocyte erythrocyte monocyte macrophage colony-forming units (CFU-GEMM) and erythroid burst forming units (BFU-E) in vitro. Conclusion : These result suggest that Brassica campestris var narinosa (BCN) and Canavalia gladiata DC have hematopoietic enhancement via hematopoietic cytokine-mediated JAK2/GATA-1/STAT-5a/b pathway, and their combinational prescription (BCN+CGD) has superior hematopoietic enhancement to those of individual extracts.

• Radiation Oncology Journal
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• v.23 no.4
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• pp.211-216
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• 2005

Purpose: It has been reported that all-trans retinoic acid (ATRA) can inhibit glioma growing in vitro. However, clinical trials with ATRA alone in gliomas revealed modest results. ATRA has been shown to increase radiosensitivity in other tumor types, so combining radiation and ATRA would be one of alternatives to increase therapeutic efficacy in malignant gliomas. Thus, we intended to know the role of catalase, which is induced by ATRA, for radiosensitivity if radiation-reduced reactive oxygen species (ROS) is removed by catalase, the effect of radiation will be reduced. Materials and Methods: A rat glioma cell line (36B10) was used for this study. The change of catalase activity and radiosensitivity by ATRA, with or without 3-amino-1, 2, 4-triazole (ATZ), a chemical inhibitor of catalase were measured. Catalase activity was measured by the decomposition of $H_2O_2$ spectrophotometrically Radiosensitivity was measured with clonogenic assay. Also ROS was measured using a 2, 7-dichlorofluorescein diacetate spectrophotometrically. Results: When 36B10 cells were exposed to 10, 25 and $50{\mu}M$ of ATRA for 48 h, the expression of catalase activity were increased with increasing concentration and incubation time of ATRA. Catalase activity was decreased with increasing the concentration of AT (1, $10{\mu}M$) dose-dependently. ROS was increased with ATRA and it was augmented with the combination of ATRA and radiation. ATZ decreased ROS production and increased cell survival in combination of ATRA and radiation despite the reduction of catalase. Conclusion: The increase of ROS is one of the reasons for the increased radiosensitivity in combination with ATRA. The catalase that is induced by ATRA doesn't decrease ROS production and radiosensitivity.

• Radiation Oncology Journal
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• v.25 no.3
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• pp.185-191
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• 2007

Purpose: This study was performed to investigate the effects of immune cell activation and the antitumor effect for the combination of treatment with X-irradiation and E/eutherococcus senticosus Maxim Root (ESMR) on mouse tumor cells. Materials and Methods: ESMR (250g) was extracted with 80% methanol, concentrated under decompression and lyophilized. To determine whether ESMR is able to activate the immune cells or not, the proliferation of splenocytes in vitro and the number of B cells and T cells in splenic lymphocytes in ESMR-pretreated mice were evaluated. X-irradiation was given to the mouse fibrosarcoma tumor cells (FSa II) by 250 kv X-irradiation machine. The cytotoxicity of ESMR was evaluated from its ability to reduce the clonogenecity of FSa II cells. In X-irradiation alone group, each 2, 4, 6 and 8 Gy was given to FSa II cells. In X-irradiation with ESMR group, 0.2 mg/ml of ESMR was exposed to FSa II cells for 1 hour before X-irradiation. Results: The proliferation of cultured mouse splenocytes and thymocytes were enhanced by the addition of ESMR in vitro. The number of B cells and T cells in mouse splenic lymphocytes was significantly increased in ESMR pretreated mice in vivo. In FSa II cells that received a combination of 0.2 mg/ml of ESMR with X-irradiation exposure, the survival fraction with a dose of 2, 4 and 6 Gy was $0.39{\pm}0.005$, $0.22{\pm}0.005$ and $0.06{\pm}0.007$, respectively. For FSa II cells treated with X-irradiation alone, the survival fraction with a dose of 2, 4 and 6 Gy was $0.76{\pm}0.02$, $0.47{\pm}0.008$ and $0.37{\pm}0.01$. The difference in the survival fraction of the mouse FSa II cells treated with and without ESMR was statistically significant (p<0.05). Conclusion: Treatment with ESMR increased cell viability of mouse splenocytes in vitro and especially the subpopulation of B cells and T cells in splenocytes in ESMR-pretreated mice. However, treatment with ESMR did not increase the level of Th and Tc subpopulations in the thymocytes. Treatment with the combination of ESMR and X-irradiation was more cytotoxic to mouse tumor cells than treatment with X-irradiation alone; this finding was statistically significant.

• Tuberculosis and Respiratory Diseases
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• v.50 no.1
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• pp.67-75
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• 2001

Background : Two tumor suppressor genes, p53 and p16, which have different roles in controlling the cell cycle and inducing apoptosis, are frequently inactivated during carcinogenesis including lung cancer. Single tumor suppressor gene therapies using either with p53 or p16 have been studied extensively. However, there is a paucity of reports regarding a combined gene therapy using these two genes. Methods : The combined effect of p53 and p16 gene transfer by the adenoviral vector on the growth of lung cancer cell lines and its interactive mechanism was investigated. Results : An isobologram showed that the co-transduction of p53 and p16 exhibited a synergistic growth in hibitory effect on NCI H358 and an additive effect on NCI H23. Cell cycle analysis demonstrated the induction of a synergistic G1/S arrest by a combined p53 and p16 transfer. This synergistic interaction was again confirmed in a soft agar confirmed in a soft agar clonogenic assay. Conclusion : These observations suggest the potential of a p53 and p16 combination gene therapy as another potent strategy in cancer gene therapy.

• Radiation Oncology Journal
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• v.19 no.3
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• pp.252-258
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• 2001

Purpose : The objectives of this study are to investigate the significance of apoptotic death compared to total cell death after $\gamma-ray$ irradiation in human H&N cancer cell lines and to find out correlation between apoptosis and radiation sensitivity. Materials and method : Head and neck cancer cell lines (PCI-1, PCI-13, and SNU-1066), leukemia cell line (CCRF-CEM), and fibroblast cell line (LM217) as a normal control were used for this study. Cells were irradiated using Cs-137 animal experiment irradiator. Total cell death was measured by clonogenic assay. Annexin-V staining was used to detect the fraction of apoptotic death. Results : Surviving fraction at 2 Gy (SF2) were 0.741, 0.544, 0.313, 0.302, and 0.100 for PCI-1, PCI-13, SNU-1066, CCRF-CEM, and LM217 cell lines, respectively. Apoptosis was detected in all cell lines. Apoptotic index reached peak value at 72 hours after irradiation in head and neck cancer cell lines, and that was at 24 hours in CCRF-CEM and LM217. Total cell death increased exponentially with increasing radiation dose from 0 Gy to 8 Gy, but the change was minimal in apoptotic index. Apoptotic fractions at 2 Gy were $46\%,\;48\%,\;46\%,\;24\%,\;and\;19\%$ and at 6 Gy were $20\%,\;33\%,\;35\%,\;17\%,\;and\;20\%$ for PCI-1, PCI-13, SNU-1066, CCRF-CEM, and LM217, respectively. The radioresistant cell lines showed more higher apoptotic fraction at 2 Gy, but there was not such correlation at 6 Gy. Conclusion : All cell lines used in this study showed apoptosis after irradiation, but time course of apoptosis was different from that of leukemia cell line and normal fibroblast cell line. Reproductive cell death was more important mode of cell death than apoptotic death in all cell lines used in this study. But there was correlation between apoptotic fraction and radiation sensitivity at 2 Gy.

• Radiation Oncology Journal
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• v.18 no.2
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• pp.133-137
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• 2000

Purpose :To investigate whether combined beta-carotene with X-Irradiation has more enhanced radition response than X-irradiation or not, we peformed a experiment about in vitro cytotoxlcity of beta-carotene and/or X-irradiation in the fibrosarcoma cells, tumor growth delay of combined beta-caroten with/or X-irradiation in the mouse fibrosarcoma. Materials and Methods : 2$\%$ emulsion of beta-carotene was serially diluted and used. X-Irradiation was given by 6 MeV linear accelerator. The cytotoxicity of beta-carotene in vitro was evaluated from clonogenic assay. To compare the cytotoxiclty between combined beta-carotene with X-irradiation and X-irradiation group, 2 mg/ml of beta-carotene was contacted to fibrosarcoma (FSall) cells for 1 hour before X-irradiation. For the tumor growth delay, single 20 Gy was given to FSall tumor hearing C3H/N mice whic was classified as beta-crotene with X-irradiation group (n=5) and X-irradiation alone group (n=5). 0.2 ml of 20 mg/kg of beta-carotene were i.p. injected to mice 30 minute before X-irradiation in the beta-crotene with X-irradiation group. The tumor growth delay defined as the time which reach to 1,000 mm$^{3}$ of tumor volume. Results : (1) Cytotoxicity in vitro: 1) survival fraction at beta-carotene concentration of 0.002,0.02,0.2 and 2 mg/ml were 0.69$\pm$0.07, 0.59$\pm$0.08, 0.08$\pm$0.008 and 0.02$\pm$0.006, respectively. 2) each survival fraction at 2, 4, 6 and 8 Gy in the 2 mg/ml of beta-carotene + X-irradiation group were 0.13$\pm$0.05, 0.03$\pm$0.005, 0.01 $\pm$0.002 and 0.009$\pm$0.0008, respectively. But each survival fraction at same irradiation dose in the X-irradiation group were 0.66$\pm$0.05, 0.40$\pm$0.04, 0.11$\pm$0.01 and 0.03$\pm$0.006, respectively(p<0.05). (2) The time which reach to 1,000 mm$^{3}$ of tumor volume of beta-carotene + X-irradiation group and X-irradiation alone group were 18, 19 days, respectively(p>0.05) Conclusion : The contact of beta-caroten to Fsall cells showed mild cytotoxicity which 띤as increased according to concentration. The cytotoxicity of combined beta-carotene with X-irradiation more increased than that of X-irradiation, additionally, And there was significant difference of cytotoxicity between two groups. But there were no significant difference of the growth delay of fibrosarcoma between two groups.