• 한국응용약물학회:학술대회논문집
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• 한국응용약물학회 2003년도 Annual Meeting of KSAP : International Symposium on Pharmaceutical and Biomedical Sciences on Obesity
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• pp.88-88
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• 2003

Cytochrome P-450 3A4 (CYP3A4) is major enzyme in human liver, the role of this is detoxification and metabolizing more than 50% clinical drugs in use. Expression of CYP3A4 is transciptionally regulated by the Pregnenolone X receptor (PXR), of which human form is Steroid and Xenobiotics receptor (SXR). SXR is activated by wide range of endogenous and exogenous compounds, and then induces CYP3A4 gene expression. In the previous study, it has been known that proximal promoter (-864 to +64) does not response to chemical inducers such as pregnenolone 16a-carbonitrile (PCN), Rifampicin, Estrogen in terms of transcription of CYP 3A4 in cultured cells. Here, we developed luciferase reporter gene assay system to detect SXR-based CYP 3A4 transcriptional activity. We have used CYP3A4-Luc plasmid that contains proximal promoter of human CYP3A4 gene upstream of the luciferase gene. We did transient transfection of 3A4-luciferase gene and SXR. In the HepG2 cells transfected with CYP3A4-Luc, when rifampicin treatment was combined with histone deacetylase inhibitor (HDAC Inhibitor), such as Trichostatin A, Hc-toxin and IN 2001 of the luciferase activity was induced 10-20 fold over control.

• 한국환경독성학회:학술대회논문집
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• 한국환경독성학회 2003년도 추계국제학술대회
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• pp.178-178
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• 2003

Cytochrome P-450 3A4 (CYP3A4) is major enzyme in human liver, the role of this Is detoxification and metabolizing more than 50% clinical drugs in use. Expression of CYP3A4 is transciptionally regulated by the Pregnenolone X receptor (PXR), of which human form is Steroid and Xenobiotics receptor (SXR). SXR is activated by wide range of endogenous and exogenous compounds, and then induces CYP3A4 gene expression. In the previous study, it has been known that proximal promoter (-864 to ＋64) does not response to chemical inducers such as pregnenolone 16a-carbonitrile (PCN), Rifampicin, Estrogen in terms of transcription of CYP 3A4 in cultured cells. Here, we developed luciferase reporter gene assay system to detect SXR-based CYP 3A4 transcriptional activity. We have used CYP3A4-Luc plasmid that contains proximal promoter of human CYP3A4 gene upstream of the luciferase gene. We did transient transfection of 3A4-luciferase gene and SXR. In the HepG2 cells transfected with CYP3A4-Luc, when rifampicin treatment was combined with histone deacetylase inhibitor (HDAC Inhibitor), such as Trichostatin A, Hc-toxin and IN 2001 of the luciferase activity was induced 10-20 fold over control.

• Molecules and Cells
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• 제20권1호
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• pp.136-141
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• 2005

Mild stresses such as high temperature ($30^{\circ}C$) or a low $H_2O_2$ concentration induced transient cell cycle arrest at G1/S or G2/M depending on the cell cycle stage at which the stress was applied. When stresses were introduced during G0 or G1, the G1/S checkpoint was mainly used; when stresses were introduced after S phase, G2/M was the primary checkpoint. The slowing of cell cycle progression was associated with transient delays in expression of A-, B-, and D-type cyclins. The delay in expression of NtcycA13, one of the A-type cyclins, was most pronounced. The levels of expression of Ntcyc29 (a cyclin B gene) and of CycD3-1 differed most depending on the applied stress, suggesting that different cellular adjustments to mild heat and a low concentration of $H_2O_2$ are reflected in the expression of these two cyclins.

• Journal of Plant Biotechnology
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• 제40권1호
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• pp.27-36
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• 2013

We isolated and functionally characterized a Dendrobium Actin1 (DmACT1) promoter that drives strong gene expression in the orchid genus Dendrobium. A genomic fragment containing the region 3227 bp upstream of the coding region of DmACT1 was obtained by inverse PCR. Detailed comparison of the full-length cDNA and genomic sequences revealed that DmACT1 has a 1374 bp first intron in the 5' UTR. However, the 5' flanking sequences upstream of the coding region showed no obvious sequence similarities compared to those of known promoters, including plant actin promoters. Serial deletion constructs of the 5' flanking region from the translation initiation codon were fused to the coding sequence of a GUS/luciferase fusion reporter to identify the regulatory elements necessary for promoter activity. Transient assays in the flowers of Dendrobium revealed that the 5' UTR-intron greatly enhanced promoter activity. Moreover, the DmACT1 promoter with its 5' UTR-intron yielded approximately 10-fold higher reporter activity than the rice Act1 promoter-intron. Our data suggest that the DmACT1 promoter with its 5' UTR-intron is a useful tool for strong expression of transgenes in Dendrobium orchids.

• 생명과학회지
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• 제15권2호
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• pp.176-181
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• 2005

Calmodulin 유전자의 발현 조절을 연구하기 위해, 벼 calmodulin promoter (RCaM-2)를 분리하여 GUS (report 유전자)에 융합하였다. GUS 활성은 정단조직, 근단 및 관다발 영역과 같은 성장조직에서 높게 발현되었다. 줄기와 페티올의 transverse 절단부위 GUS 활성은 관다발계의 안쪽에 제한되었으며 관다발계의 외부에 위치한 피층과 표피에서는 강하게 발현된 식물에서도 GUS 활성이 나타나지 않았다. GUS 활성은 어린 조직에서 특이적으로 발현되었으며 상처에 의해서 신속하게 증가하였다. RCaM-2 promoter는 세포분열이 왕성한 어린조직이나 생장점에서 강하게 발현되며 mechanical 신호에 의해서 현저히 유도되었다. 이러한 결과는 RCaM-2 유전자의 5'-flanking 영역이 상처에 의해서 유도되는 발현을 조절하는 것으로 추정된다.

• Journal of Applied Biological Chemistry
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• 제64권3호
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• pp.205-211
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• 2021

In order to test the functionality of Panax ginseng thioredoxin 1 (PgTRX1) isolated from fermented wild ginseng roots, a transient effect on physiological activity were performed over a short time frame using the Agrobacterium infiltration technique. The PgTRX1 gene isolated from fermented wild ginseng was confirmed to have a size of 579 bp, and the expression of PgTRX1 was the highest in the sample after 6 h of fermentation. As a result of constructing this gene and confirming the infiltration reaction mediated by Agrobacterium in tobacco leaves, it was found that the expression of the NbHSR203j gene was also induced as PgTRX1 expression increased. As a result of measuring the biological activity of the infiltration samples, the total phenol content increased by 35.45±1.84 to 49.01±1.84 ㎍ GAE/mL compared to the control, and the total flavonoid amount of 9.52±0.41 to 9.82±0.25 ㎍ QE/mL was slightly high. From these results, Agrobacterium-mediated PgTRX1 appears to be related to the hypersensitive response induction mechanism of plants and the production of secondary metabolites such as phenolic substances.

• Molecules and Cells
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• 제27권5호
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• pp.583-589
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• 2009

GTP cyclohydrolase I (GTPCH) is a key enzyme in the de novo synthesis of tetrahydrobiopterin. Previously, the Drosophila melanogaster GTPCH gene has been shown to be expressed from two different promoters (P1 and P2). In our study, the 5'-flanking DNA regions required for P1 and P2 promoter activities were characterized using transient expression assay. The DNA regions between -98 and +31, and between -73 and +35 are required for efficient P1 and P2 promoter activities, respectively. The regions between -98 and -56 and between -73 and -41 may contain critical elements required for the expression of GTPCH in Drosophila. By aligning the nucleotide sequences in the P1 and P2 promoter regions of the Drosophila melanogaster and Drosophila virilrs GTPCH genes, several conserved elements including palindromic sequences in the regions critical for P1 and P2 promoter activities were identified. Western blot analysis of transgenic flies transformed using P1 or P2 promoter-lacZ fusion plasmids further revealed that P1 promoter expression is restricted to the late pupae and adult developmental stages but that the P2 promoter driven expression of GTPCH is constitutive throughout fly development. In addition, X-gal staining of the embryos and imaginal discs of transgenic flies suggests that the P2 promoter is active from stage 13 of embryo and is generally active in most regions of the imaginal discs at the larval stages.

• Asian Pacific Journal of Cancer Prevention
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• 제15권18호
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• pp.7825-7829
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• 2014

Objective: To investigate the effect of high expression of XAF1 in vivo or in vitro on lung cancer cell growth and apoptosis. Methods: 1. The A549 human lung cancer cell line was transfected with Ad5/F35 - XAF1, or Ad5/F35 - Null at the same multiplicity of infection (MOI); (hereinafter referred to as transient transfected cell strain); XAF1 gene mRNA and protein expression was detected by reverse transcription polymerase chain reaction (RT-PCR) and Western blotting respectively. 2. Methyl thiazolyl tetrazolium (MTT) and annexin V-FITC/PI double staining were used to detect cell proliferation and apoptosis before and after infection of Ad5/F35 - XAF1 with Western blotting for apoptosis related proteins, caspase 3, caspase - 8 and PARP. 3. After the XAF1 gene was transfected into lung cancer A549 cells by lentiviral vectors, and selected by screening with Blasticidin, reverse transcription polymerase chain reaction (RT-PCR) and Western blotting were applied to detect mRNA and protein expression, to establish a line with a stable high expression of XAF1 (hereinafter referred to as stable expression cell strain). Twenty nude mice were randomly divided into groups A and B, 10 in each group: A549/XAF1 stable expression cell strain was subcutaneously injected in group A, and A549/Ctrl stable cell line stable expression cell strain in group B (control group), to observe transplanted tumor growth in nude mice. Results: The mRNA and protein expression of XAF1 in A549 cells transfected by Ad5/F35 - XAF1 was significantly higher than in the control group. XAF1 mediated by adenovirus vector demonstrated a dose dependent inhibition of lung cancer cell proliferation and induction of apoptosis. This was accompanied by cleavage of caspase -3, -8, -9 and PARP, suggesting activation of intrinsic or extrinsic apoptotic pathways. A cell strain of lung cancer highly expressing XAF1 was established, and this demonstrated delayed tumor growth after transplantation in vivo. Conclusion: Adenovirus mediated XAF1 gene expression could inhibit proliferation and induce apoptosis in lung cancer cells in vitro; highly stable expression of XAF1 could also significantly inhibit the growth of transplanted tumors in nude mouse, with no obvious adverse reactions observed. Therefore, the XAF1 gene could become a new target for lung cancer treatment.

• 대한한의학회지
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• 제24권2호
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• pp.81-93
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• 2003

Objectives : Recently, the new therapeutic tool, that is herbal acupuncture, has been developed and applied to various diseases including the cerebrovascular accident. The main characteristics of herbal acupuncture are a combination of acupuncture and herbal medicine. It was not well known the therapeutic effect and the mechanism of herbal acupuncture on transient forebrain ischemic injury, although it has been used frequently in clinics. The objective was to determine the effect of folium artemisiae argyi (艾葉) herbal acupuncture on the trasient forebrain ischemic injured rat. Methods : In this study, the effects of folium artemisiae argyi (艾葉) herbal acupuncture on the $LR_3$ named Taechung, on neuroprotection after the transient forebrain ischemia were investigated in Sprague-Dawely rats. Expressions of cFos, FosB and BDNF protein in the hippocampus and cortex were observed at 2 hrs and 48 hrs after transient forebrain ischemia by immunohistochemistry and ELISA technique. Results : Expression of cFos protein was increased slightly in the hippocampus and cortex at 2 hrs after transient forebrain ischemia, but FosB protein was increased highly comparing to cFos protein. However, pretreatment with folium artemisiae argyi' herbal acupuncture on $LR_3$ significantly increased expression of cFos protein and significantly decreased expression of FosB protein compared to control group. These features were observed in the retrosplenial granular cortex as well as the hippocampus. Also, pretreatment with folium artemisiae argyi' herbal acupuncture on $LR_3$ significantly increased the expression of BDNF protein in the hippocampus ($263.26{\pm}44.56{\;}pg/ml$ extracted by water, $275.35{\pm}51.47{\;}pg/ml$ extracted by moxa tar)and the cortex ($102.33{\pm}13.65{\;}pg/ml$ extracted by water, $109.54{\pm}9.37{\;}pg/ml$ extracted by moxa tar) compared to the hippocampus ($134.07{\pm}2.96{\;}pg/ml$) and the cortex ($61.16{\pm}4.11{\;}pg/ml$) in control group at 48 hrs after transient forebrain ischemia. Conclusions : These results suggest that pretreatment with folium artemisiae argyi' herbal acupuncture on $LR_3$ has neuroprotective effect on transient forebrain ischemia and the herbal acupunture on $LR_3$ may be related to antioxidative function of folium artemisiae argyi.

• Korean Journal of Acupuncture
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• 제20권3호
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• pp.61-80
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• 2003

Objectives : Acupuncture and herbal medicine have been used to prevent and treat the cerebrovascular accident, such as a stroke, and many studies of acupuncture and moxibustion concerning to the stroke have been undertaken in the human and various animals. Recently, the new therapeutic tool, that is herbal acupuncture, has been developed since the 1950' and applied to various diseases including the cerebrovascular accident. The main characteristics of herbal acupuncture are a combination of acupuncture and herbal medicine. It was not well known the therapeutic effect and the mechanism of herbal acupuncture on transient forebrain ischemic injury, although it has been used frequently in clinics. Methods : In this study, effects of folium Artemisiae Argyi and moxa tar' herbal acupuncture on the $GV_{20}$, named Baek-Hue, on neuroprotection after the transient forebrain ischemia were investigated in Sprague-Dawely rats. Expressions of cFos, FosB and BDNF protein in the hippocampus and cortex were observed at 2 hrs and 48 hrs after transient forebrain ischemia by immunohis- tochemistry and ELISA technique. Results : Expression of cFos protein was increased slightly in the hippocampus and cortex at 2 hrs after transient forebrain ischemia, but FosB protein was increased highly comparing to cFos protein. However, pretreatment with folium Artemisiae Argyi or moxa tar' herbal acupuncture on $GV_{20}$ significantly increased expression of cFos protein and significantly decreased expression of FosB protein compared to control group, respectively. These features were observed in the motor cortex and retrosplenial granular cortex as well as the hippocampus. Also, pretreatment with folium Artemisiae Argyi and moxa tar' herbal acupuncture on$GV_{20}$ significantly increased the expression of BDNF protein in the hippocampus and the cortex compared to control group at 48 hrs after transient forebrain ischemia, respectively. Conclusions : These results suggest that pretreatment with folium Artemisiae Argyi or moxa tar' herbal acupuncture on $GV_{20}$ has neuroprotective effect on transient forebrain ischemia and theherbal acupuncture on $GV_{20}$ may be related to antioxidative function.