• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• 제32권6호
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• pp.495-505
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• 2006

In this study we evaluate the effects of bFGF-BB and PDGF on in vitro proliferation, differentiation and mineralization of mesenchymal stem cells (MSCs) from rat. MSCs were prepared from the bone marrow of 6 or 7-week-old male rats with a technique previously described by Maniatopoulos et al. in 1988. Lineage differentiation to osteogenesis, chondrogenesis and adipogenesis were performed. At first, we characterized the cultured cell on passage 1, 3, 5, 7 with immunocytochemical staining using CD29, 44, 34, 45, ${\alpha}$-SMA and type I collagen. And to study the effects of bFGF and PDGF-BB on proliferation, differentiation and mineralization, we seeded the expanded cell at a density of 6 $6{\times}10^3\;cells/cm^2$ to 100-mm dish for evaluation of cell proliferation and MTT assay was carried out on day 2, 4, 7, 9. We also resuspended the cells with same density $(6{\times}10^3\;cells/cm^2)$ to 24 well plates for subculture. On the following day, the attached cells were exposed to 2.5ng/ml bFGF and/or 25ng/ml PDGF-BB daily during 5 days. The osteocalcin (OC) level was assessed and mineral contents were evaluated with alizarin red S staining on subculture day 2, 7, 14, 21. We identified the mesenchymal stem cell from the bone marrow derived cells of rat through their successful multi-differentiation and stable display of its phenotype. And bFGF and PDGF-BB showed the effect that inhibited osteoblastic differentiation and mineralization mildly in above concentration at in vitro culture. This study was supported by grant 04-2004-0120 from the Seoul National University Hospital Research Fund.

• 한국자원식물학회:학술대회논문집
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• 한국자원식물학회 2019년도 춘계학술대회
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• pp.33-33
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• 2019

The AKT signaling pathway is a highly regulated cell signaling system that forms a network with other cell signaling pathways. Hence, the AKT signaling pathway mediates several important cellular functions that include cell survival, proliferation, cell migration, and et cetera. Irregularities that led overactive AKT signaling have been linked to many diseases such as cancer and metabolic-associated diseases. Hence, modulating the overactive AKT signaling pathway via inhibitor is a tantalizing prospect for treatment of cancer and metabolic-associated diseases. Two inhibitors of the AKT signaling pathway will be presented in this symposium: 1) Bisleuconothine A (BisA), a bisindole alkaloid that inhibit autophagy and 2) Ceramicine B (CerB), a limonoid that inhibit adipogenesis. The first topic is on a bisindole alkaloid, BisA and its mechanism in inducing autophagosome formation in lung cancer cell line, A549.(1) Since most autophagy inducing agents generally induce apoptosis, we found that BisA does not induce apoptosis even in high dose. BisA up-regulation of LC3 lipidation is achieved through mTOR inactivation. The phosphorylation of PRAS40, a mTOR repressor was suppressed by BisA. This observation suggested that BisA inactivates mTOR via suppression of PRAS40 phosphorylation. Interestingly, the phosphorylation of AKT, an upstream regulator of PRAS40 phosphorylation was also down-regulated by BisA. These findings suggested that Bis-A induces autophagosomes formation by interfering with the AKT-mTOR signaling pathway. The second topic is on CerB and its mechanism in inhibiting adipogenesis in preadipocytes cell line, MC3T3-G2/PA6.(2,3) CerB inhibits the phosphorylation of protein kinase B (AKT) at the Thr308 position but not the Ser473. Consequently, the phosphorylation of FOXO3 which is located downstream of AKT is also inhibited. Considering that FOXO3 is an important regulator of PPARγ which is a key factor in adipogenesis, CerB may inhibit adipogenesis via the AKT-FOXO3 signaling pathway. Taken together, both BisA and CerB highlighted the potential of AKT signaling pathway modulation as an approach to induce autophagy and inhibit the formation of fat cells, respectively.

• Nutrition Research and Practice
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• 제7권2호
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• pp.89-95
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• 2013

Dietary inorganic sulfur is the minor component in our diet, but some studies suggested that inorganic sulfur is maybe effective to treat cancer related illness. Therefore, this study aims to examine the effects of inorganic sulfur on cell proliferation and gene expression in MDA-MB-231 human breast cancer cells. MDA-MB-231 cells were cultured the absence or presence of various concentrations (12.5, 25, or 50 ${\mu}mol/L$) of inorganic sulfur. Inorganic sulfur significantly decreased proliferation after 72 h of incubation (P < 0.05). The protein expression of $ErbB_2$ and its active form, $pErbB_2$, were significantly reduced at inorganic sulfur concentrations of 50 ${\mu}mol/L$ and greater than 25 ${\mu}mol/L$, respectively (P < 0.05). The mRNA expression of $ErbB_2$ was significantly reduced at an inorganic sulfur concentration of 50 ${\mu}mol/L$ (P < 0.05). The protein expression of $ErbB_3$ and its active form, $pErbB_3$, and the mRNA expression of $pErbB_3$ were significantly reduced at inorganic sulfur concentrations greater than 25 ${\mu}mol/L$ (P < 0.05). The protein and mRNA expression of Akt were significantly reduced at an inorganic sulfur concentration of 50 ${\mu}mol/L$ (P < 0.05), but pAkt was not affected by inorganic sulfur treatment. The protein and mRNA expression of Bax were significantly increased with the addition of inorganic sulfur concentration of 50 ${\mu}mol/L$ (P < 0.05). In conclusion, cell proliferation was suppressed by inorganic sulfur treatment through the ErbB-Akt pathway in MDA-MB-231 cells.

• 대한화장품학회지
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• 제27권1호
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• pp.17-27
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• 2001

A cell-based assay system for monitoring NF-$textsc{k}$B activity was developed to determine the influence of activated NF-$textsc{k}$B in human HaCaT cells. The pNF-$textsc{k}$B-SEAP-NPT plasmid that permits expression of the secreted alkaline phosphatase (SEAP) reported gene in response to the NF-$textsc{k}$B activity and contains neomycin phosphotransferase (NPT) gene for the geneticin resistance in host cells was constructed and transfected into human keratinocyte cell line HaCaT. Human HaCaT transfectant cells secreted the SEAP enzyme into the culture medium in a time-dependent manner until 72h. NF-$textsc{k}$B activities were measured in the SEAP reporter gene assay using a fluorescent detection method. The treatment of HaCaT cell transfectants with known antioxidants [e.g., N-acetyl-L-cysteine and vitamin C] showed inhibition of NF-$textsc{k}$B activity in a time-and concentration-dependent manner. The phorbol 12-myristate 13-acetate (PMA) known as a stimulator of NF-$textsc{k}$B expression demonstrated that it increased NF-$textsc{k}$B activity in a time- and concentration-dependent manner. This assay system could be used to determine the quantitative measurement of NF-$textsc{k}$B activity in the human skin and allow the screening of anti-inflammatory agents from various synthetic chemicals and natural products for dermatological purpose. Abbrevitions used: NF-$textsc{k}$B, nuclear factor kappa B; I-$textsc{k}$B, Inhibitory kappa B; SEAP, secreted alkaline phosphatase; NPT, neomycin phosphotransferease; PCR, polymerase chain reaction: dNTP, deoxynucleoside triphosphates; DMEM, dulbecco’s modified eagle medium; FBS, fetal bovine serum; PBs, phosphate-buffered saline; MUP, 4-methylumbellifery phosphate; NAC, N-acetyl-L-cysteine; DMSO, dimethyl sulfoxide; PMA, phorbol 12-myristate 13-acetate.

• The Korean Journal of Physiology and Pharmacology
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• 제4권6호
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• pp.507-513
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• 2000

ErbB3/HER3 is a cell surface receptor which belongs to the ErbB/HER subfamily of receptor protein tyrosine kinases. When expressed in NIH/3T3 cells, ErbB3 can form heterodimeric coreceptor with endogenous ErbB2. Among known intracellular effectors of the ErbB2/ErbB3 are mitogen-activated protein kinase (MAPK) and phosphoinositide (PI) 3-kinase. In the present study, we studied relative contributions of above two distinct signaling pathways to the heregulin-induced mitogenic response via activated ErbB3. For this, clonal NIH-3T3 cell lines expressing wild-type ErbB3 and ErbB3 mutants were stimulated with $heregulin{\beta}_1$. While cyclin D1 level was markedly high and further increased by treatment of heregulin in cells expressing wild-type ErbB3, the elimination of either Shc binding or PI 3-kinase binding lowered both levels. This result was supported by the reduction of cyclin $D_1$ expression by preteatment with MAPK kinase inhibitor or PI 3-kinase inhibitor before stimulation with heregulin. In accordance with the cyclin $D_1$ expression, elimination of either Shc binding or PI 3-kinase binding reduced the heregulin-induced DNA synthesis and cell growth rate. Our results obtained by the comparison of wild-type and ErbB3 mutants indicate that the full induction of the cell cycle progression through $G_1/S$ phase by ErbB3 activation is dependent on both Shc/MAPK and PI 3-kinase signal transduction pathways.

• BMB Reports
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• 제44권8호
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• pp.523-528
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• 2011

To identify novel genes that are regulated by promoter methylation, a combinational approach involving in silico mining followed by molecular assay was performed. From the expression microarray data registered in the European bioinformatics institute (EBI), genes showing downregulation in breast cancer cells were initially screened and then selected by e-Northern analysis using the Unigene database. A series of these in silico methods identified CAMK2B and ARFGEF1 as candidates, and the two genes were revealed to be hypermethylated in breast cancer cell lines and hypomethylated in normal breast cell lines. Additionally, cancer cell lines showed downregulated expression of these genes. Furthermore, treatment of the cancer cell lines with a demethylation agent, 5-Aza-2'-deoxycytidine, recovered expression of CAMK2B and ARFGEF1, implying that hypermethyaltion silenced gene activity in cancer cells. Taken together, promoter methylations of CAMK2B and ARFGEF1 are novel epigenetic markers identified in breast cancer cell lines and can be utilized for the application to clinical cancer tissues.

• Journal of Microbiology and Biotechnology
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• 제19권10호
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• pp.1122-1126
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• 2009

The exogenously added cadaverine is effective in protecting Vibrio vulnificus from methyl viologen (MV)-induced superoxide stress at pH 8.5. Such a protective effect by cadaverine was not observed at pH 7.5. Consistently, the accumulated level of intracellular cadaverine at pH 8.5 is approximately four times as much as that of the control cell at pH 7.5. Cadaverine accumulation is not affected by MV. The protection of V. vulnificus by cadaverine from superoxide stress was abolished when cadB coding for the lysine-cadaverine antiporter was interrupted. However, the cadaverine-mediated protection was complemented with cadB DNA. Therefore, CadB of V. vulnificus not only acts as a lysine-cadaverine antiporter at acid pH to neutralize the external medium, but also mediates cadaverine uptake at alkaline pH to result in cell protection from superoxide stress.

• Journal of Microbiology and Biotechnology
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• 제16권8호
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• pp.1192-1200
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• 2006

An apple polygalacturonase-inhibiting protein (PGIP), which specifically inhibits endopolygalacturonase (PG, EC 3.2.1.15) from Botryosphaeria dothidea, was purified from Botryosphaeria dothidea-infected apple (Malus domestica cv. Fuji) fruits. The purified apple PGIP had a molecular mass of 40 kDa. The N-terminal amino acid sequence of the purified protein showed high homologies to those of PGIP from pear (100%), tomato (70%), and bean (65%). We also purified polygalacturonase (PG) from B. dothidea. The PG hydrolyzes pectic components of plant cell walls. When the extracted apple pectic cell wall material was treated with purified apple PGIP and B. dothidea PG, the amount of uronic acid released was lower than that treated with B. dothidea PG alone. This result demonstrates that PGIP functions specifically by inhibiting cell wall maceration of B. dothidea PG Furthermore, we characterized the de novo function of the PGIP against PG on the solubilization and depolymerization of polyuronides from cell wall of apple fruits inoculated with B. dothidea. This result demonstrated that the PGIP of plants exhibits one of the direct defense mechanisms against pathogen attack by inhibiting PGs that are released from pathogens for hydrolysis of cell wall components of plants.

• BMB Reports
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• 제30권4호
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• pp.269-273
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• 1997

Heptocvte-specific expression induced by Hepatitis B virus (HBV) enhancer 2-core gene promoter was examined in various hepatocyte and non-hepatocyte cell lines. using non-viral and retroviral vector systems in which chloramphenicol acetyltransferase (CAT) is used as a reporter. The non-viral plasmid containing the HBV enhancer 2-core promoter exhibited 22 and 66% of CAT activities in hepatoma cell lines. HepG2 and Hep3B, respectively when compared with CAT activity expressed by CMV promoter. The CAT activities, however. were found to be marginal in other tested hepatoma cell lines as well as mouse primary hepatocytes and non-hepatocytes. The HBV enhancer 2 located upstream the CMV promoter did not affect the CMV promoter activity nor provided hepatocyte-specific expression. Transfection of retroviral plasmid DNA containing the HBV enhancer 2-core promoter as an internal promoter exhibited high and specific CAT expression in HepG2 and Hep3B cell lines but the activity value was 5 to 10 fold lower than the non-viral plasmid with identical promoter. These results suggest that the usage of HBV enhancer 2-core promoter for liver specific expression is limited to certain vectors and hepatocyte cell lines.

• 한국통신학회논문지
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• 제19권5호
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• pp.842-856
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• 1994

광대역 종합정보 통신망 (B-ISDN)이 수용할 여러 서비스들은 다양한 지연, 지연 변화 및 셀 손실 확률 요구 사항을 가지고 있다. B-ISDN을 위한 적절한 제어 방법의 설계는 매우 중요하고 어려운 문제이다. 본 논문에서는, 그러한 다양한 요구사항을 만족하기 위하여, 지연 및 손실 우선 순위 모두를 가진 다종화 알고리즘을 제안한다. 손실 우선 순위의 구현을 위하여, 음성 셀은 폐기 불가한 셀(즉, 높은 우선 순위의 셀) 및 폐기 가능한 셀(즉, 낮은 우선 순위의 셀)로 생성됨을 가정하였다. 낮은 순위의 음성셀은 통신망내에서 혼잡이 발생하면 폐기될 수 있다. 본 셀 탈락 방법은 음성 및 데이터의 지연뿐만 아니라 셀 손실도 감소 시키는 것을 보여주고 있다. 이러한 부하 감소 방법은 광대역 종합정보 통신망의 이용율을 크게 개선 시킬 수 있을 것으로 기대된다.