• Bulletin of the Korean Chemical Society
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• 제31권9호
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• pp.2607-2612
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• 2010

Human tissue-type plasminogen activator (tPA) is a valuable thrombolytic agent used to successfully treat acute myocardial infarction, thromboembolic stroke, peripheral arterial occlusion, and venous thromboembolism. Recombinant tPA is accumulated as an inactive form in inclusion bodies of E. coli and is refolded in vitro, which is accompanied by extensive aggregation. In the present study, a tPA protease domain was expressed in an active soluble form in the cytosol of E. coli Rosetta-gami cells, which allowed disulfide bond formation and supplied the tRNA molecules required for six rarely used codons in E. coli. This strategy increased the amount of soluble protease domain protein and avoided the cumbersome refolding process. The purified protease domain not only degraded tPA substrate peptides but also formed a covalently bound complex with plasminogen activator inhibitor-1, as does full-length tPA. Soluble expression and purification of tPA domains may aid in functional analyses of this multi-domain protein, which has been implicated in many physiological and pathological processes.

• Tuberculosis and Respiratory Diseases
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• 제70권6호
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• pp.462-473
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• 2011

Background: Smoking is a risk factor for idiopathic pulmonary fibrosis (IPF), but the mechanism of the association remains obscure. There is evidence demonstrating that plasminogen activator inhibitor-1 (PAI-1) is involved in the progression of pulmonary fibrosis. This study was to determine whether the administration of small interfering RNA (siRNA) targeting PAI-1 or PAI-1 inhibitor to the cigarette smoking extract (CSE)-exposed rat alveolar type II epithelial cells (ATII cells) limits the epithelial-mesenchymal transition (EMT). Methods: ATII cells were isolated from lung of SD-rat using percoll gradient method and cultured with 5% CSE. The EMT was determined from the ATII cells by measuring the real-time RT PCR and western blotting after the PAI-1 siRNA transfection to the cells and after administration of tiplaxtinin, an inhibitor of PAI-1. The effect of PAI-1 inhibitor was also evaluated in the bleomycin-induced rats. Results: PAI-1 was overexpressed in the smoking exposed ATII cells and was directly associated with EMT. The EMT from the ATII cells was suppressed by PAI-1 siRNA transfection or administration of tiplaxtinin. Signaling pathways for EMT by smoking extract were through the phosphorylation of SMAD2 and ERK1/2, and finally Snail expression. Tiplaxtinin also suppressed the pulmonary fibrosis and PAI-1 expression in the bleomycin-induced rats. Conclusion: Our data shows that CSE induces rat ATII cells to undergo EMT by PAI-1 via SMAD2-ERK1/2-Snail activation. This suppression of EMT by PAI-1 siRNA transfection or PAI-1 inhibitor in primary type II alveolar epithelial cells might be involved in the attenuation of bleomycin-induced pulmonary fibrosis in rats.

• Biomolecules & Therapeutics
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• 제21권2호
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• pp.107-113
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• 2013

Plasminogen activator inhibitor-1 (PAI-1) is a member of serine protease inhibitor family, which regulates the activity of tissue plasminogen activator (tPA). In CNS, tPA/PAI-1 activity is involved in the regulation of a variety of cellular processes such as neuronal development, synaptic plasticity and cell survival. To gain a more insights into the regulatory mechanism modulating tPA/PAI-1 activity in brain, we investigated the effects of proteasome inhibitors on tPA/PAI-1 expression and activity in rat primary astrocytes, the major cell type expressing both tPA and PAI-1. We found that submicromolar concentration of MG132, a cell permeable peptide-aldehyde inhibitor of ubiquitin proteasome pathway selectively upregulates PAI-1 expression. Upregulation of PAI-1 mRNA as well as increased PAI-1 promoter reporter activity suggested that MG132 transcriptionally increased PAI-1 expression. The induction of PAI-1 downregulated tPA activity in rat primary astrocytes. Another proteasome inhibitor lactacystin similarly increased the expression of PAI-1 in rat primary astrocytes. MG132 activated MAPK pathways as well as PI3K/Akt pathways. Inhibitors of these signaling pathways reduced MG132-mediated upregulation of PAI-1 in varying degrees and most prominent effects were observed with SB203580, a p38 MAPK pathway inhibitor. The regulation of tPA/PAI-1 activity by proteasome inhibitor in rat primary astrocytes may underlie the observed CNS effects of MG132 such as neuroprotection.

• Reproductive and Developmental Biology
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• 제33권4호
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• pp.203-209
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• 2009

The present study was performed to identify changes of plasminogen activator (PA) and plasminogen activator inhibitor (PAI) in porcine oviduct epithelial cells (POECs) during the estrous cycle. POECs obtained from ovary in pre-ovulatory (Pre-Ov), early to mid-luteal stage (Early-mid L) and post-ovulatory stage (Post-Ov). For the examine of PA activity, $1{\times}10^5$ fresh cells of POECs were cultured in DMEM/Ham F-12 containing 10% FBS and 0.2% amphotericin under humidified atmosphere of 5% $CO_2$ in air and $38^{\circ}C$. The urokinase-type PA (uPA) was observed at 7 days of POECs culture. PA activity was measured with culture prolonged of 0, 3, 6, 12 and 24 h after culture of 7 days. The PA activity were high significantly (p<0.05) at 12 h of culture, but PA activity were decreased with culture periods increased. The PA activity in POECs of Post-Ov stage were higher significantly (p<0.05) than that of Early-mid L and Pre-Ov stage. When PAI-1 and PAI-2 were added during the POECs culture, the PA were observed significant low activity (p<0.05). The PA activity and protein expression were decreased by PA inhibitor. This results suggest that PAI-1 and PAI-2 have a suppressive action on change of PA activity during the estrous cycle of pigs. Specifically, this study using PA inhibitor was effect the PA activity and PAI expression in oviduct epithelial cells in pigs.

• Childhood Kidney Diseases
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• 제8권1호
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• pp.26-32
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• 2004

목 적 : 소아 신증후군 환자에서 과응고성 경향을 가지고 있으며 Plasminogen activator inhibitor-1(PAI-1)은 강력하게 섬유소의 용해를 감소시키는 당단백으로 최근 몇몇 연구에 의하면 신증후군에서 증가된 PAI-1과 사구체내의 섬유소원이나 섬유소와 관련된 항체의 침착이 연관되어 있음을 시사하고 있다. 저자는 PAI-1 유전자 다형성 중에서 A-844 G 대립유전자 다형성의 빈도와 유전형을 소아의 신증후군에서 임상경과와의 연관성을 비교 평가하였다. 방 법 : 2001년 10월부터 2003년 1월까지 경희대학교 부속병원 동서신장병연구소에 방문한 146명의 신증후군 환아와 249명의 대조군을 대상으로 하였고 신증훈군 환아는 infrequent relapser(IR)와 frequent relapser(FR)로 다시 나누었다. 이들에 대해 PAI-1 promoter gene의 A-844 G에 대한 중합효소 연쇄반응-제한효소절편길이 다형현상(PCR-RFLP)을 이용하여 유전자형을 A/A, A/G, G/G로 분류하여 비교 분석하였다. 통계는 Graph Pad Prism 통계분석 소프트웨어 version 2.0을 사용하였고 95% 신뢰구간과 P value는 0.05보다 작은 것을 의미 있게 보았다. 결 과 : PAI-1 promoter gene의 A- 844G 다형성의 분포는 대조군애서 G/G 81(32.5%), A/A 42(16.9%), G/A 126(50.6%)이였고 신증후군 그룹 중 IR 그룹은 G/G 29(34.1%), A/A 15(17.7%), G/A 41(48.2%)이였으며 FR 그룹에서는 G/G 17(27.9%), A/A 18(29.5%), G/A 26(42.6%)이었다. 단지 PAI-1 gene의 A-844G의 다형성중 A/A 유전자형이 신증후군 환아 중 FR군에서만 대조군에 비하여 유의하게 증가하였다(16.9% vs 29.5%, OR=2.06, P=0.0251). 반면 A/G 유전자형(OR=0.73, P=0.2639)이나 G/G 유전자형(OR=0.80, P=0.4828)은 통계학적인 의의가 없었고 IR군에서 대조군에 비하여 A/A(OR=1.06, P=0.8690), A/G(OR=0.91, P=0.7063), G/G(OR=1.07, P=0.7880)으로 모두 통계학적으로 유의하지 않았다. 결 론 : A-844G AA 유전자형을 가진 신증후군 환자와 FR와 통계적인 상관관계가 있음을 알수 있었다. 이는 향후 PAI-1 유전자와 과응고성과 관련된 생화학적 검사와의 연관성에 대한 추가적인 조사와 특히, 신증후군의 과응고성과 신증후군 재발과의 연관성이 있는지에 관한 연구가 더 필요할 것으로 사료된다.

• 생명과학회지
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• 제19권10호
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• pp.1438-1443
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• 2009

이 연구는 신장섬유아 세포를 이용하여 LPS에 의해 유도된 신장섬유화 표적유전자인 plasminogen activator inhibitor (PAI-1) 발현과 Ascofuranone (AF)에 의한 신장섬유화 저해효과를 연구하였다. 이 연구를 통해 LPS가 PAI-1의 발현을 농도 및 시간 의존적으로 증가시켜 LPS가 신장섬유화 유도물질임을 확인 할 수 있었다. 또한 LPS로 유도된 PAI-1 mRNA 및 단백질 발현 레벨이 AF에 의해 저해되었으며, 신장섬유화의 또 다른 대표유전자인 fibronectin의 단백질 발현도 AF에 의해 억제되어 AF가 신장섬유화를 저해하는 사실을 확인할 수 있었다. 그리고 AF에 대한 PAI-1 프로모터 활성을 조사하기 위하여 p800-PAI-1-luc을 신장섬유아 세포에 형질전환 시킨 결과, AF가 PAI-1의 전사 활성 조절을 통해 발현을 억제한다는 것을 확인하였다. ERK-1/2의 상위에 존재하는 MEK inhibitor를 처리하여 PAI-1의 발현을 확인한 결과에서도 AF를 처리한 경우와 동일하게 PAI-1 발현이 저해되어 LPS로 유도된 PAI-1의 발현이 ERK-1/2에 의해 조절됨을 알 수 있었다. 또한 LPS로 유도된 ERK-1/2의 인산화가 AF 농도의존적으로 저해된 결과는, AF가 ERK-1/2의 활성저해를 통하여 PAI-1 발현을 조절한다는 사실을 확인 할 수 있었다. 따라서 이러한 연구결과 AF가 신장섬유화를 저해하는 유력한 후보물질로서의 가능성을 제시하였다.

• Molecular & Cellular Toxicology
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• 제2권3호
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• pp.212-215
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• 2006

Several reports have suggested a possible relationship between blood coagulation factors and schizophrenia. Plasminogen activator inhibitor type 1 (PAI-1) belongs to a serine protease inhibitor family, which regulates fibrinolysis and proteolysis by inhibiting plasminogen activation. The purpose of this study was to investigate the association of polymorphisms of the PAI-1 gene with schizophrenia in Korean population. Two important polymorphisms (-675 4G/5G and -844 G/A) located on promoter region of the PAI-1 gene were analyzed on 178 schizophrenia patients and 226 controls. The genotypic and allelic associations of -675 4G/5G were found significant. Furthermore, haplotype analysis revealed significant result, which suggests that -675 4G/5G polymorphism might confer increased susceptibility for schizophrenia in Korean population.

• Tuberculosis and Respiratory Diseases
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• 제75권4호
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• pp.140-149
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• 2013

Background: Plasminogen activator inhibitor type 1 (PAI-1), an important regulator of plasminogen activator system which controls degradation of extracellular membrane and progression of tumor cells, and PAI-1 gene polymorphic variants have been known as the prognostic biomarkers of non-small cell lung cancer patients. Recently, experimental in vitro study revealed that transforming growth factor-${\beta}1$ initiated PAI-1 transcription through epithelial growth factor receptor (EGFR) signaling pathway. However, there is little clinical evidence on the association between PAI-1 A15T gene polymorphism and prognosis of Korean population with pulmonary adenocarcinoma and the influence of activating mutation of EGFR kinase domain. Methods: We retrospectively reviewed the medical records of 171 patients who were diagnosed with pulmonary adenocarcinoma and undergone EGFR mutation analysis from 1995 through 2009. Results: In all patients with pulmonary adenocarcinoma, there was no significant association between PAI-1 A15T polymorphic variants and prognosis for overall survival. However, further subgroup analysis showed that the group with AG/AA genotype had a shorter 3-year survival time than the group with GG genotype in patients with EGFR mutant-type pulmonary adenocarcinoma (mean survival time, 24.9 months vs. 32.5 months, respectively; p=0.015). In multivariate analysis of 3-year survival for patients with pulmonary adenocarcinoma harboring mutant-type EGFR, the AG/AA genotype carriers had poorer prognosis than the GG genotype carriers (hazard ratio, 7.729; 95% confidence interval, 1.414-42.250; p=0.018). Conclusion: According to our study of Korean population with pulmonary adenocarcinoma, AG/AA genotype of PAI-1 A15T would be a significant predictor of poor short-term survival in patients with pulmonary adenocarcinoma harboring mutant-type EGFR.

• Nutrition Research and Practice
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• 제14권5호
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• pp.463-477
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• 2020

BACKGROUND/OBJECTIVES: Many studies have suggested that Rhus verniciflua Stokes (RVS) and its extract are anticancer agents. However, RVS had limited use because it contains urushiol, an allergenic toxin. By improving an existing allergen-removal extraction method, we developed a new allergen-free Rhus verniciflua Stokes extract (RVSE) with higher flavonoid content. In this study, we examined whether RVSE inhibits the ability of AGS gastric cancer cells to migrate and invade. MATERIALS/METHODS: The flavonoids content of RVSE was analyzed by HPLC. The effects of RVSE on migration and invasion in AGS cells were analyzed by each assay kit. Matrix metalloproteinase (MMP)-9, plasminogen activator inhibitor-1 (PAI-1) and urokinase-type plasminogen activator (uPA) protein expression was analyzed by protein antibody array. The Phosphorylation of signal transducer and activator of transcription (STAT) 3 were assayed by Western blot analysis. RESULTS: RVSE treatment with 0-100 ㎍/mL dose-dependently reduced the ability of AGS cells to migrate and invade. Notably, treatment with RVSE strongly inhibited the expression of MMP-9 and uPA and the phosphorylation of STAT3. In contrast, RVSE treatment dramatically increased the expression of PAI-1. These results indicate that the inhibition of MMP-9 and uPA expression and STAT3 phosphorylation and the stimulation of PAI-1 expression contributed to the decreased migration and invasion of AGS cells treated with RVSE. CONCLUSIONS: These results suggest that RVSE may be used as a natural herbal agent to reduce gastric cancer metastasis.

• BMB Reports
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• 제47권7호
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• pp.376-381
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• 2014

Enzymatic oxidation of pyrogallol was efficiently transformed to an oxidative product, purpurogallin (PPG). Here, the anticoagulant activities of PPG were examined by monitoring activated partial thromboplastin time (aPTT), prothrombin time (PT), and the activities of thrombin and activated factor X (FXa). And, the effects of PPG on expression of plasminogen activator inhibitor type 1 (PAI-1) and tissue-type plasminogen activator (t-PA) were evaluated in tumor necrosis factor (TNF)-${\alpha}$ activated human umbilical vein endothelial cells (HUVECs). Treatment with PPG resulted in prolonged aPTT and PT and inhibition of the activities of thrombin and FXa, as well as inhibited production of thrombin and FXa in HUVECs. In addition, PPG inhibited thrombin-catalyzed fibrin polymerization and platelet aggregation. PPG also elicited anticoagulant effects in mice. In addition, treatment with PPG resulted in significant reduction of the PAI-1 to t-PA ratio. Collectively, PPG possesses antithrombotic activities and offers a basis for development of a novel anticoagulant.