Kim, Seong-A;Woo, Hong-Jung;Kim, Young-Chul;Lee, Jang-Hoon
The Journal of Internal Korean Medicine
/
v.29
no.1
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pp.177-188
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2008
Objectives : This study was performed to investigate the anti-fibrogenic effect of Artemisiae Capillaris Herba on cultured rat hepatic stellate cells. Materials and Methods : Hepatic stellate cells(HSC-T6) were treated with various concentrations of Artemisiae Capillaris Herba extract for 24 hours. The extraction was done either with distilled water or 50% EtOH. After the treatment, cell viability, proliferation, procollagen levels and the mRNA of the collagen type 1a2 and ASMA were measured by using MTT assay, BrdU assay, RT-PCR, and Procollagen Type I C-peptide EIA Kit. Results : The viability and proliferation of the hepatic stellate cells were decreased as the concentration increased. The mRNA expression decreased consistently with the volume of the secreted procollagen with the extraction made with distilled water, which indicates the herb has inhibitory effect on fibrogenesis of the liver by regulating one of the fibrosis associated genes in transcription. However, it increased in 50% EtOH extraction, which shows that a more stable reaction is expected of the extraction made with distilled water than the extraction made with 50% EtOH. The production of procollagen was decreased by a low-concentration treatment with Artemisiae Capillaris Herba, but increased by a high concentration. It seemed that the cells were responding to Artemisiae Capillaris Herba in low- concentrations, thus producing small amounts of collagen. When the drug was administered at high enough concentration to give direct toxicity to cells, the ability of cells to produce collagen was activated, and the overproduction of collagen was observed as an undesirable results. Conclusion : These results suggest that Artemisiae Capillaris Herba is beneficial in the treatment of cirrhotic patients as well as for the patients with chronic hepatitis when extracted with water in the proper concentrations.
Cytochrome P450 1B1 (CYP1B1) acts as a hydroxylase for estrogen and activates potential carcinogens. Moreover, its expression in tumor tissues is much higher than that in normal tissues. Despite this association between CYP1B1 and cancer, the detailed molecular mechanism of CYP1B1 on cancer progression in HeLa cells remains unknown. Previous reports indicated that the mRNA expression level of Herc5, an E3 ligase for ISGylation, is promoted by CYP1B1 suppression using specific small interfering RNA, and that ISGylation may be involved in ubiquitination related to ${\beta}-catenin$ degradation. With this background, we investigated the relationships among CYP1B1, Herc5, and ${\beta}-catenin$. RT-PCR and western blot analyses showed that CYP1B1 overexpression induced and CYP1B1 inhibition reduced, respectively, the expression of $Wnt/{\beta}-catenin$ signaling target genes including ${\beta}-catenin$ and cyclin D1. Moreover, HeLa cells were treated with the CYP1B1 inducer $7,12-dimethylbenz[{\alpha}]anthracene$ (DMBA) or the CYP1B1 specific inhibitor, tetramethoxystilbene (TMS) and consequently DMBA increased and TMS decreased ${\beta}-catenin$ and cyclin D1 expression, respectively. To determine the correlation between CYP1B1 expression and ISGylation, the expression of ISG15, a ubiquitin-like protein, was detected following CYP1B1 regulation, which revealed that CYP1B1 may inhibit ISGylation through suppression of ISG15 expression. In addition, the mRNA and protein expression levels of Herc5 were strongly suppressed by CYP1B1. Finally, an immunoprecipitation assay revealed a direct physical interaction between Herc5 and ${\beta}-catenin$ in HeLa cells. In conclusion, these data suggest that CYP1B1 may activate $Wnt/{\beta}-catenin$ signaling through stabilization of ${\beta}-catenin$ protein from Herc5-mediated ISGylation for proteosomal degradation.
Bae, Yong Chan;Park, Suk Young;Nam, Su Bong;Herh, Jae Young;Kang, Young Seok
Archives of Plastic Surgery
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v.33
no.1
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pp.5-12
/
2006
Protein tyrosine kinase(PTK), protein kinase C(PKC), oxidase, as a mediator, take a significant role in signal transduction pathway of angiogenesis. The authors utilized the inhibitors, targeting the formation of three co-enzyme in signal transduction pathway in order to quantify the suppression of abnormal vascular endothelial cell proliferation induced by DMH, to compare the level suppression in each up-regulated growth factors, CTGF, CYR61, $ITG{\beta}1$, FHL2, and to identify the relationship between abnormal cell proliferation and signal transduction pathway. Five groups were established; Control group, Group of DMH, Group of DMH-mixed Herbimycin, inhibitor of protein tyrosine kinase, Group of DMH-mixed Calphostin C, inhibitor of protein kinase C, Group Of Dmh-Mixed 10U Catalase, Inhibitor Of oxidase. The rise of vascular endothelial cell was compared by MTT assay, and four growth factors were analysed with RT-PCR method, at pre-administration, 4, 8, 12, and 24 hours after administration. In comparison of abnormal proliferation of vascular endothelial cell induced by DMH, suppression was noticed in Herbimycin and Calphostin C group, and Calphostin C group revealed higher suppression effect. Nevertheless, Catalase group did not have any suppression. In manifestation of four growth factors, Herbimycin and Calphostin C group presented similar manifestation with control group, except in $ITG{\beta}$. Catalse group had similar manifestation with DMH group in all four growth factors. Abnormal proliferation of vascular endothelial cell induced by DMH have a direct relationship with PTK and PKC, more specifically to PKC. Oxidase was confirmed not to have any relevance.
The Journal of the Korean Society for Microbiology
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v.35
no.2
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pp.181-190
/
2000
To investigate resistance to lamivudine (3TC), we examined the incidence of M184V in 20 HIV-1 patients treated with 3TC for $13.1{\pm}9$ months. Fourteen of 20 patients had been exposed to zidovudine (ZDV) or didanosine (ddI) prior to 3TC therapy. Nested PCR targeting to reverse transcriptase (RT) and direct sequencing were performed for peripheral blood mononuclear cells sampled serially. There were resistance mutations to ZDV in at least 9 patients at baseline, although there was no resistance mutation to 3TC. We could detect M184V in 6 (30%) out of 20 patients. The incidence of M184V increased as the duration of therapy prolongs (13% in samples <12 months; 47% in samples ${\ge}12$ months). The frequency of mutation M184V was higher in patients with previous mutation to ZDV than in patients with wild type. Resistance mutation was not detected in 7 patients. This study shows that resistance to 3TC tends to develop rapidly in patients with baseline mutations or two drugs combination therapy than in those treated simultaneously with triple drugs. This report is the first on resistance to 3TC in Korean AIDS patients.
In agro-ecosystems worldwide, some of the most important and devastating diseases are caused by soil-borne necrotrophic fungal pathogens, against which crop plants generally lack genetic resistance. However, plants have evolved approaches to protect themselves against pathogens by stimulating and supporting specific groups of beneficial microorganisms that have the ability to protect either by direct inhibition of the pathogen or by inducing resistance mechanisms in the plant. One of the best examples of protection of plant roots by antagonistic microbes occurs in soils that are suppressive to take-all disease of wheat. Take-all, caused by Gaeumannomyces graminis var. tritici, is the most economically important root disease of wheat worldwide. Take-all decline (TAD) is the spontaneous decline in incidence and severity of disease after a severe outbreak of take-all during continuous wheat or barley monoculture. TAD occurs worldwide, and in the United States and The Netherlands it results from a build-up of populations of 2,4-diacetylphloroglucinol (2,4-DAPG)-producing fluorescent Pseudomonas spp. during wheat monoculture. The antibiotic 2,4-DAPG has a broad spectrum of activity and is especially active against the take-all pathogen. Based on genotype analysis by repetitive sequence-based-PCR analysis and restriction fragment length polymorphism of phlD, a key 2,4-DAPG biosynthesis gene, at least 22 genotypes of 2,4-DAPG producing fluorescent Pseudomonas spp. have been described worldwide. In this review, we provide an overview of G. graminis var. tritici, the take-all disease, Pseudomonas biocontrol agents, and mechanism of disease suppression.
Rapid advances in pharmacogenomic research have provided important information to improve drug selection, to maximize drug efficacy, and to minimize drug adverse reaction. The SNPs that are the most abundant type of genetic variants have been proven as valid biomarkers to give information on the prediction of pharmacokinetic/pharmacodynamic properties of drugs based on genotype. In order to elucidate a correlation between SNPs of calcium channel encoding gene and adverse reactions of calcium channel blockers, we investigated SNPs in CACNA1C gene known as a binding site of calcium channel blocker. 96 patients with hypertension who had taken or are taking an antihypertensive drug, 1,4-dihydropyridine (DHP) were included for analysis. These patients were composed of 47 patients with adverse drug reactions (ADR) such as edema from calcium channel blockers and 49 patients without ADR as a control group. The exons encoding the drug binding sites were amplified by PCR using specific primers, and SNPs were analyzed by direct sequencing. We found that there was no SNP in the exons encoding DHP binding site, but four novel SNPs in the exon-intron junction region. However, four novel SNPs were not associated with the ADR of calcium channel blockers. In conclusion, this study showed that ADR from calcium channel blockers may not be caused by SNPs of the binding sites of calcium channel blockers in CACNA1C gene.
Kim, Yun-Jung;Park, Eun-Hye;Kim, Gyu-Dong;Nam, Kyoungphile;Chung, Tai Hak
Journal of Korean Society on Water Environment
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v.20
no.5
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pp.506-511
/
2004
Maintaining dissolved oxygen (DO) at sufficiently low concentration in the aeration tank at a wastewater treatment plant (WWTP) is essential for reduction of the costs of operation and maintenance. On the other hand, the low DO level may result in adverse effect on the integrity of the activated sludge, A typical and disastrous outcome frequently experienced is the outgrowth of filamentous microorganisms, which is called as filamentous bulking, In addition to the traditional methods such as sludge settleability and microscopic observation of the culture, molecular techniques including polymerase chain reaction (PCR) amplification followed by 16S rRNA sequencing were applied to identify filamentous bacteria present in bulking sludge under a condition of low DO concentration, Two morphologically distinct groups, presumably consisting of Sphaerofilus nafans, and Eikelboom Type 1701 or Type 1851, were identified through microscopic observation. They were further confirmed by subsequent 16S rRNA sequencing. Dominant filamentous bacteria identified by the molecular techniques were consisted of three major groups. Sequences of partial 16S rRNA cloned showed that the filamentous bulking organisms were closely related to Eikelboom Type 021N and Eikelboom Type 1701, and Sphaerotilus natans, respectively. Molecular methods were found to possess a strong potential of direct examination of the microbial community of an activated sludge system.
Direct cell-cell communication through connexin (Cx) complexes is a way to achieve functional accordance of cells within a tissue or an organ. The initial segment (IS), a part of the epididymis, plays important roles in sperm maturation. Steroid hormones influence on expression of a number of genes in the IS of adult animals. However, developmental effect of sex hormones on the gene expression in the IS has not been examined. In this study, estradiol benzoate (EB, an estrogen agonist) or flutamide (Flu, an androgen antagonist) was exogenously administrated at 1 week of postnatal age, and expressional changes of Cx genes in the IS were determined at 4 months of age by a quantitative real-time PCR analysis. Treatment of EB at $0.015{\mu}g/kg$ body weight (BW) increased expression of Cx30.3, 31.1, and 43 genes. However, treatment of 1.5 mg EB/kg BW resulted in expressional decreases of Cx31, 32, and 45 genes and caused increases of Cx30.3 and 43 gene expression. Significant decreases of Cx31, 31.1, 32, 37, and 45 gene expression were detected with a treatment of $500{\mu}g\;Flu/kg$ BW, while expression of Cx43 gene was significantly increased with a treatment of $500{\mu}g\;Flu/kg$ BW. A treatment of $50{\mu}g\;Flu/kg$ BW led to significant increases of Cx30.3, 32, 37, 40, and 43 gene expression. These findings imply that exogenous exposure of steroidal hormones during the early developmental period would result in aberrant expression of Cx genes in the adult IS.
Cell-cell direct communication through channel-forming molecules, connexin (Cx), is essential for a tissue to exchange signaling molecules between neighboring cells and establish unique functional characteristics during postnatal development. The corpus epididymis is a well-known androgen-responsive tissue and involves in proper sperm maturation. In the present research, it was attempted to determine if expression of Cx isoforms in the corpus epididymis in the adult is modulated by exposure to estrogenic or anti-androgenic compound during the early postnatal period. The neonatal male rats at 7 days of age were subcutaneously injected by estradiol benzoate (EB) at low-dose ($0.015{\mu}g/kg$ body weight) or high-dose ($1.5{\mu}g/kg$ body weight) or flutamide (Flu) at low-dose ($500{\mu}g/kg$ body weight) or high-dose (50 mg/kg body weight). The corpus epididymis collected at 4 months of age was subjected to evaluate expressional changes of Cx isoforms by quantitative real-time PCR. Treatment of low-dose EB resulted in increases of Cx32, Cx37, and Cx45 transcript levels, while exposure to high-dose EB decreased expression of Cx26, Cx30.3, Cx31, Cx31.1, Cx32, Cx40, Cx43, and Cx45. Treatments of Flu caused significant decreases of expression of all examined Cx isoforms, except Cx37 and Cx43 shown no expressional change with high-dose Flu treatment. These findings imply that expression of most Cx isoforms present in the corpus epididymis would be transcriptionally regulated by actions of androgen and/or estrogen during postnatal period.
The present study was designed to obtain porcine growth hormone binding protein (pGHBP) improved biological activation as derived mutation in binding site with growth horlnone (GH). A 756 bp of fragment encoding the extracellular domain of pGHBP gene was cloned from the total RNA of porcine fat tissue by reverse transcriptase polymerase chain reaction (RT-PCR) and created mutation in positions 26 and 122 using site-directed mutagenesis method. Position 26 is one and it is near to get on five potential N-linked glycosylation sites located in the extracellular domain of porcine growth hormone receptor known to have a direct influence on combination with GH. Position 122 is known as one of conformational epitope in bovine. It was over-expressed in E. coli using pET-32(c) expression vector and precisely purified by S-protein agarose and enterokinase. In our results, we was obtained pmGHBP of 30 kDa. It suggests to study the effects of the pmGHBP on cell proliferation in vitro and growth rate in vivo after administration.
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