• Proceedings of the Korean Vacuum Society Conference
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• 2011.02a
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• pp.20-20
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• 2011

It has been known that, under certain conditions, application of low-temperature atmospheric-pressure plasmas can enhance proliferation of cells. In this study, conditions for optimal cell proliferation were examined for various cells relevant for orthopaedic applications. Plasmas used in our experiments were generated by dielectric barrier discharge (DBD) with a helium flow (of approximately 3 litter/min) into ambient air at atmospheric pressure by a 10 kV~20 kHz power supply. Such plasmas were directly applied to a medium, in which cells of interest were cultured. The cells examined in this study were human synoviocytes, rat mesenchymal stem cells derived from bone marrow or adipose tissue, a mouse osteoblastic cell line (MC3T3-E1), a mouse embryonic mesenchymal cell line (C3H-10T1/2), human osteosarcoma cells (HOS), a mouse myoblast cell line (C2C12), and rat Schwann cells. Since cell proliferation can be enhanced even if the cells are not directly exposed to plasmas but cultured in a medium that is pre-treated by plasma application, it is surmised that long-life free radicals generated in the medium by plasma application stimulate cell proliferation if their densities are appropriate. The level of free radical generation in the medium was examined by dROMs tests and correlation between cell proliferation and oxidative stress was observed. Other applications of plasma medicine in orthopaedics, such as plasma modification of artificial bones and wound healing effects by direct plasma application for mouse models, will be also discussed. The work has been done in collaboration with Prof. H. Yoshikawa and his group members at the School of Medicine, Osaka University.

• Journal of Acupuncture Research
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• v.20 no.3
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• pp.63-74
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• 2003

목적 : 한의학에서 관절염이나 진통치료에 사용되어 왔던 봉독약침액이 인간 골육종 세포주인 MG-63 세포에서 항종양효과가 있는지 연구하고자 한다. 특히 본 실험에서는 이러한 봉독의 종양발생 억제작용이 세포사멸과 관련이 있는지, 그리고 프로스타글란딘 합성 효소인 cyclooxygenase(COX)-2의 억제와 관련이 있는지를 연구하고자 한다. 방법 : 인간 골육종 세포주에서 세포사멸의 변화를 관찰하기 위해서 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium brimide(MTT) assay, 4,6-diamidino-2-phenylindole (DAPI), DNA fragmentation assay 및 reverse transcription-polymerase chain reaction(RT-PCR) 방법을 이용하였다. 결과 : 세포독성 검사에서 봉독은 MG-63 세포에서 농도-의존적으로 세포독성을 나타내었다. 이러한 봉독의 세포독성이 세포사멸로 인한 것인지를 여러 가지 형태로 검사한 결과 봉독에 의한 세포독성은 TUNEL 검사와 DAPI 염색시 세포사멸의 특징적인 소견들을 나타내었고, flow cytometric 분석에서도 세포사멸을 의미하는 세포주기의 변화들을 나타내었다. 봉독이 COX-2의 발현에 미치는 영향을 RT-PCR로 실험한 결과 봉독은 COX-2 mRNA의 발현을 선택적으로 억제하였다. 결론 : 본 실험의 결과 봉독은 COX-2 mRNA의 발현을 억제함으로써 골육종 세포에서 세포사멸을 유발하고 그 결과 항종양효과를 나타내는 것으로 보여진다.

• KSBB Journal
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• v.13 no.4
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• pp.357-363
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• 1998

The antitumor effects of mice fed with cell lysate of Lactobacillus plantarum were studied. The abdominal cancer induced by Sarcoma-180 was markedly inhibited and the expected life span was extended by 60% for the Balb/c mice fed with L. plantarum cell lysate for two weeks. A similar result was obtained for the rat inoculated with Spontaneous Osteosarcoma(SOS). The primary tumor volume of SOS was reduced by 70% for the rats fed with L. plantarum cell lysate (100mg/kg/day) for one week before the inoculation of SOS, while only 42% for the rats fed with the same amount of cell lysate for one week after the inoculation of tumor cell line, SOS. As lung was the metastasis site of SOS, the weight of lung was measured to determine the degree of metastasis inhibition by the L. plantarum cell lysate feeding. The rats fed with cell lysate for one week showed a remarkable inhibition of lung metastasis by 63%(before) and 46%(after), respectively. These results indicate that the feeding of L. plantarum cell lysate to mouse or rat can induce a strong stimulation of mucosal or systemic immune system and these effects results in an efficient antitumor activity.

• Proceedings of the Ginseng society Conference
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• 2002.10a
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• pp.376-384
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• 2002

Object To find out which of the 27 ginsenosides isolated from Panax ginseng C.A. Mey that may inhibit the proliferation of human osteosaocoma cell line $U_2OS$. Methods Effects of each individual ginsenoside on the proliferation of $U_2OS$ cell were studied by determining the viability of cancer cells during culture with or without the presence of the test compound. DNA assay was determined by flow cytometry. Results Ginsonosides -Ro, $-Rh_l,\;-Rh_2,\;-F_1\;and\;-L_8$ at concentrations of 5 ,umol/L could obviously suppress the proliferation of $U_2OS$ cells while ginsenosides $-Rg_1,\;-F_3,$ -Rf, PPT and PT significantly inhibited the cancer cells. Flow cytometry revealed that ginsenosides $-Ro,-Rg_1-Rf,-F_1-Rh_2,PPT$ and PT induced cell cycle arrest at $G_0/G_1$ phase with obvious decrease of cell count at Sand $G_2+M$ phase, Moreover, ginsenosides $-Rf_1,-Rg_1,\;-F_1$ and PPT induced significantly high rates of cell death as compared with the control. Conclusion These data suggested that ginsenosides inhibited $U_2OS$ proliferation Via cell cycle arrest or induction of cell death.

• Journal of Periodontal and Implant Science
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• v.35 no.2
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• pp.345-357
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• 2005

Prostaglandin plays a significant role in the local control of bone metabolism associated with periodontal disease. ${\Delta}^{12}-PGJ_2$ is a natural $PGD_2$ metabolite that is formed in vivo in the presence of plasma. It is known for ${\Delta}^{12}-PGJ_2$ to stimulate calcification in osteoblastic cells. Bone morphogenetic protein(BMP) stimulated osteoblastic differentiation in various types of cells and greatly enhanced healing of bony defects. The purpose of this study was to evaluate the effect of rhEMP-2 on ${\Delta}^{12}-PGJ_2$ induced osteoblastic differentiation and mineralization in vitro. A human osteosarcoma cells line Saos-2 were cultured. In the test groups, 10-7M of ${\Delta}^{12}-PGJ_2$ or mixture of 10-8M of ${\Delta}^{12}-PGJ_2$ and 100ng/ml of rhBMP-2 or 100ng/ml of rhEMP-2 were added to culture media. After 1 day, 2 days and 4 days of culture period, the cell number was measured. Alkaline phosphatase activity was measure at 3 days. Reverse transcription polymerase chain reaction(RT-PCR) was performed to determine the expression of mRNA of bone matrix protein at 8 hours, 1 day and 7 days. The ability to produce mineralized nodules in rat osteoblasts(MC3T3-E1) was evaluated at 21 days. The results were as follows : 1. rhEMP-2 or mixture of rhBMP-2 and ${\Delta}^{12}-PGJ_2$ inhibited cell proliferation of human osteosarcoma cells. 2. rhEMP-2 or mixture of rhBMP-2 and ${\Delta}^{12}-PGJ_2$ stimulated alkaline phosphatase activity significantly higher than ${\Delta}^{12}-PGJ_2$ alone. 3. rhBMP-2 or mixture of rhEMP-2 and ${\Delta}^{12}-PGJ_2$ stimulated mineralization compared to ${\Delta}^{12}-PGJ_2$ alone. 4. mRNA of alkaline phosphatase, BMP-2, cbfa 1, Type I collagen were detected in the group treated with ${\Delta}^{12}-PGJ_2$/rhBMP-2, rhBMP-2 alone, ${\Delta}^{12}-PGJ_2$ alone. These results show that mixture of ${\Delta}^{12}-PGJ_2$ and rhBMP-2 causes more bone formation than ${\Delta}^{12}-PGJ_2$ alone while the bone formation effects of mixture of ${\Delta}^{12}-PGJ_2$ and rhBMP-2 are less than those of rhBMP-2 alone. Further researches would be necessary to clarify the interactions of these agents.

• BMB Reports
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• v.55 no.12
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• pp.627-632
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• 2022

Dickkopf-1 (DKK1) is a secreted protein that acts as an antagonist of the canonical WNT/β-catenin pathway, which regulates osteoblast differentiation. However, the role of DKK1 on osteoblast differentiation has not yet been fully clarified. Here, we investigate the functional role of DKK1 on osteoblast differentiation. Primary osteoprogenitor cells were isolated from human spinal bone tissues. To examine the role of DKK1 in osteoblast differentiation, we manipulated the expression of DKK1, and the cells were differentiated into mature osteoblasts. DKK1 overexpression in osteoprogenitor cells promoted matrix mineralization of osteoblast differentiation but did not promote matrix maturation. DKK1 increased Ca⁺ influx and activation of the Ca⁺/calmodulin-dependent protein kinase II Alpha (CAMK2A)-cAMP response element-binding protein 1 (CREB1) and increased translocation of p-CREB1 into the nucleus. In contrast, stable DKK1 knockdown in human osteosarcoma cell line SaOS2 exhibited reduced nuclear translocation of p-CREB1 and matrix mineralization. Overall, we suggest that manipulating DKK1 regulates the matrix mineralization of osteoblasts by Ca⁺-CAMK2A-CREB1, and DKK1 is a crucial gene for bone mineralization of osteoblasts.

• Molecular & Cellular Toxicology
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• v.3 no.1
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• pp.36-45
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• 2007

Several features of the implant surface, such as topography, roughness, and composition play a relevant role in implant integration with bone. This study was conducted in order to determine the effects of different-coatings on Ti surfaces on the biological responses of a human osteoblast-like cell line (MG63). MG63 cells were cultured on HA (Hydroxyapatite coating on Titanium), Ano (HA coating on anodized surface Titanium), Zr (zirconium-coating on Titanium), and control (non-coating on Titanium). The morphology of these cells was assessed by SEM. The cDNAs prepared from the total RNAs of the MG63 were hybridized into a human cDNA microarray (1,152 elements). The appearances of the surfaces observed by SEM were different on each of the three dental substrate types. MG63 cells cultured on HA, Ano, Zr, and control exhibited cell-matrix interactions. In the expression of several genes were up-, and down-regulated on the different surfaces. The attachment and expression of key osteogenic regulatory genes were enhanced by the surface morphology of the dental materials used.

• Journal of Life Science
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• v.15 no.6 s.73
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• pp.994-1004
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• 2005

The dihydrofolate reductase (dhfr) promoter contains cis-acting element for the transcription factors Spl and E2F. Transcription of dhfr gene shows maximal activity during the Gl/S phase of cell cycle. The member of the Spl transcriptional factor family can act as both negative and positive regulators of gene expression. There was a report that Spl-Rb and E2F4-pl30 complexes cooperate to establish stable repression of dhfr gene expression in CHOC400 cells. Here, we examined the role of HDAC in dhfr, cyclin E, and cyclin A gene regulation using the histone deacetylation inhibitor, trichostatin A (TSA) in U2OS and C33A cells, a Rb-positive human osteosarcoma cell line, and a Rb-negative cervical carcinoma cell line, respectively. When the dhfr promoter constructs were applied in U2OS cells, TSA markedly stimulated over 14-fold of dhfr promoter activity through dhfr-Spl sites by the deletion of an E2F element. In contrast, the deletion of dhfr-Spl binding sites completely abolished promoter stimulation by TSA. The dhfr promoter activity including dhfr-Spl sites increased only 2-fold in C33A cells. Promoter activity containing only dhfr-E2F site did not have much effect by the treatment of TSA in both U2OS and C33A cells. On the other hand, treatment with TSA induced significantly mRNA expression of dhfr and cyclin E, whereas levels of cyclin A decreased in U2OS cells, but had no effect in C33A cells. These results indicate that TSA have contradictory effect, activation of dhfr and cyclin E genes on Gl phase, and down-regulation of cyclin A on G2 phase through transcriptional regulation in U2OS cells.

• Journal of Nutrition and Health
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• v.53 no.2
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• pp.111-120
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• 2020

Purpose: Corosolic acid (CA), also known as 2α-hydroxyursolic acid, is present in numerous plants, and is reported to exhibit anti-cancer and anti-proliferative activities in various cancer cells such as osteosarcoma, hepatocellular carcinoma, lung adenocarcinoma, and colon cancer. However, the anti-cancer activity of CA on human breast cancer cells and the underlying mechanisms remain to be elucidated. The present study aimed to investigate the anticancer effects of CA in the human breast cancer cell line, MDA-MB-231. Methods: Cell viability, reactive oxygen species (ROS) production, apoptosis marker protein expression, migration, invasion rate, and vascular endothelial growth factor (VEGF) levels were assessed by treating MDA-MB-231 cells to increasing concentrations of CA. Results: The results showed that CA significantly inhibited the cell proliferation of MDA-MB-231 cells in a dose-dependent manner. To assess the effect of CA on apoptosis, nuclei of MDA-MB-231 cells were stained with DAPI solution. Chromatin condensation, which indicates apoptosis, was observed to increase dose-dependently. In addition, western-blot analysis revealed elevated levels of the apoptosis marker proteins (Bax and cleaved caspase 3) subsequent to MDA-MB-231 exposure to CA. ROS production was also increased in the CA-induced apoptosis in MDA-MB-231 treated cells. Interestingly, CA exposure resulted in significantly decreased migration and invasion rates in the MDA-MB-231 cells. Data further revealed that exposure to CA markedly decreased the VEGF concentration, thereby contributing to a reduction in angiogenesis. Conclusion: Our results determined that exposure to CA induces anti-proliferation, apoptosis, and ROS production, and suppresses cell migration and invasion rate in MDA-MB-231 cells. Taken together, these results indicate the potential of CA to be applied as an effective chemotherapeutic agent for treating breast cancer.

• The Journal of Pediatrics of Korean Medicine
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• v.16 no.1
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• pp.39-74
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• 2002

Recently, increased attention has been paid to the growth of the height of children and adolescents. To accelerate growth, velvet antlers are typically used in Oriental medicine. The present study investigated the effects of velvet antlers of velvet antlers on bone growth using the cell line of Human Osteosarcoma (Hos), derived from the bone-generating cells essential to bone growth. In order to give certain stress to this Hos, the medium contained 1% FBS was used for culturing for Hos cell instead of 10% in control. In this condition of which the proliferation had been significantly decreased, the ethanol extract of upper part of velvet antlers was added, As a result, the cells proliferation rate was significantly increased. Using Oligonucleotide DNA microarray, comparison and analyses were done to see what kind of specific genes would be differentially expressed. The result showed that as opposed to the control group, the stressed group indicated a decrease in the expressions of 6 kinds of genes such as, Id1, retinoid X receptor(RXRB) and 14-3-3 epsilon, etc. The velvet antler treated group, as opposed to the control group, showed a decreased in the expressions of 8 kinds of genes such as Id1, etc. and an increase in the expressions of 24 kinds of genes. The number of genes that showed differences in the velvet antler treated group compared with the stressed group was 7 the expression of 1 kind of gene was decreased, and the expressions of 6 kinds of genes were increased. Considering the mechanism by which velvet antlers affected the growth of osteoblast through reviewing the functions of these genes, the following results were attained. The constraint in the proliferation of Hos cells resulting from the medium contained 1% FBS seems to be caused by three important factors: 1) the decrease of the expression of 14-3-3 epsilon involved in the signal transduction and metabolism of growth, 2) the decrease of the expression of Id1 gene involved in the metabolism of bone formation, and 3) the decreased of expression of RXRB gene involved in the metabolism of retinoci acid. It is suggested that the improvement of the cell proliferating effects by velvet antler treatment, in stressed condition si mediated by increment of 6 genes particularly 14-3-3 epsilon, RXRB, and IGF2, with are the crucial factors for the cell growth and differentiation, metabolism of retinoic acid and osteoblast proliferation, respectively.