• Journal of Microbiology and Biotechnology
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• 제21권12호
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• pp.1299-1305
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• 2011

An important modification of thrombolytic agents is resistance to plasminogen activator inhibitor-1 (PAI-1). In previous studies, a new truncated PAI-1-resistant variant was developed based on deletion of the first three domains in t-PA and the substitution of KHRR 128-131 amino acids with AAAA in the truncated t-PA. The novel variant expressed in a static culture system of Chinese Hamster Ovary (CHO) DG44 cells exhibited a higher resistance to PAI-1 when compared with the full-length commercial drug; Actylase. In the present study, the truncated-mutant protein was expressed in CHO DG44 cells in 50 ml orbital shaking bioreactors. The final yield of the truncated-mutant in the culture was 752 IU/ml, representing a 63% increase compared with the static culture system. Therefore, these results suggest that using the combined features of a transient and stable expression system is feasible for the production of novel recombinant proteins in the quantities needed for preclinical studies.

• BMB Reports
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• 제40권2호
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• pp.180-188
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• 2007

In vivo studies have demonstrated that aldosterone is an independent contributor to glomerulosclerosis. In the present study, we have investigated whether aldosterone itself mediated glomerulosclerosis, as angiotensin II (Ang II) did, by inducing cultured renal mesangial cells to produce plasminogen activator inhibitor-1 (PAI-1), and whether these effects were mediated by aldosterone-induced increase in transforming growth factor $\beta_1$ (TGF-$\beta_1$) expression and cellular reactive oxygen species (ROS) activity. Quiescent rat mesangial cells were treated by aldosterone alone or by combination of aldosterone and spironolactone, Ang II, neutralizing antibody to TGF-$\beta_1$ or antioxidant Nacetylcysteme (NAC). This study indicate that aldosterone can increase PAI-1 mRNA and protein expression by cultured mesangial cells alone, which is independent of aldosterone-induced increases in TGF-$\beta_1$ expression and cellular ROS. The effects on PAI-1, TGF-$\beta_1$ and ROS generation were markedly attenuated by spironolactone, a mineralocorticoid receptor antagonist, which demonstrate that mineralocorticoid receptor (MR) may play a role in mediating these effects of aldosterone.

• The Korean Journal of Physiology and Pharmacology
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• 제14권6호
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• pp.385-390
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• 2010

Excessive extracellular matrix (ECM) accumulation is the main feature of chronic renal disease including diabetic nephropathy. Plasminogen activator inhibitor (PAI)-1 is known to play an important role in renal ECM accumulation in part through suppression of plasmin generation and matrix metalloproteinase (MMP) activation. The present study examined the effect of PAI-1 antisense oligodeoxynucleotide (ODN) on fibronectin upregulation and plasmin/MMP suppression in primary mesangial cells cultured under high glucose (HG) or transforming growth factor (TGF)-${\beta}1$, major mediators of diabetic renal ECM accumulation. Growth arrested and synchronized rat primary mesangial cells were transfected with $1\;{\mu}M$ phosphorothioate-modified antisense or control mis-match ODN for 24 hours with cationic liposome and then stimulated with 30 mM D-glucose or 2 ng/ml TGF-${\beta}1$. PAl-1 or fibronectin protein was measured by Western blot analysis. Plasmin activity was determined using a synthetic fluorometric plasmin substrate and MMP-2 activity analyzed using zymography. HG and TGF-${\beta}1$ significantly increased PAI-1 and fibronectin protein expression as well as decreased plasmin and MMP-2 activity. Transient transfection of mesangial cells with PAI-1 antisense ODN, but not mis-match ODN, effectively reversed basal as well as HG- and TGF-${\beta}1$-induced suppression of plasmin and MMP-2 activity. Both basal and upregulated fibronectin secretion were also inhibited by PAI-1 antisense ODN. These data confirm that PAI-1 plays an important role in ECM accumulation in diabetic mesangium through suppression of protease activity and suggest that PAI-1 antisense ODN would be an effective therapeutic strategy for prevention of renal fibrosis including diabetic nephropathy.

• 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.

• Journal of Chest Surgery
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• 제32권4호
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• pp.333-340
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• 1999

배경: PAI-1은 t-PA의 억제인자로서 섬유소융해계에 작용을 하여 혈전형성을 유발한다. PAI-1은 동맥경화된 혈관벽에서 분비가 된다. PAI-1의 증가는 동맥경화증의 위험인자가 되는 당뇨병과 고혈압이 동반된 환자에서 보이며 혈전증유발에 위험인자가 될 수 있다. 본 연구는 고혈당과 인슐린 및 angiotensin II가 PAI-1의 생성 및 평활근세포의 증식에 미치는 영향을 규명하고자 하였다. 대상 및 방법:흰쥐 대동맥평활근세포를 5.5 mM과 22 mM의 포도당 배양액을 사용하여 배양하였다. 배양액에 angiotensin II 및 인슐린을 농도 및 배양시간에 따라 첨가하여 Northern blotting방법으로 PAI-1 유전자발현을 나타내었다. 또한 세포 증식에 대한 포도당, 인슐린 및 angiotensin II의 영향을 규명하기 위하여 MTT assay를 사용하였다. 결과: 5.5 mM과 22 mM의 포도당 배양액에서 angiotensin II(100 nM)를 첨가하여 배양한 결과, 22 mM 포도당 배양액에서 PAI-1 mRNA 발현이 증가되었으며 angiotensin II 투여 4시간에 최고치에 도달하였고 6시간까지 지속되었다. 5.5 mM, 22 mM의 포도당 배양액에 angiotensin II의 농도를 0, 10, 100, 200 nM 투여하여 배양한 결과, PAI-1 mRNA의 발현은 angiotensin II 농도에 따른 증가를 보였으며 22 mM 포도당 배양액시 더욱 뚜렷하게 증가되었다. 배양액에 angiotensin II(100 nM)과 인슐린(100 nM)을 투여하여 배양한 결과, PAI-1 mRNA의 발현은 angiotensin II 단독으로 투여시 증가하였으나 인슐린을 첨가하였을 때는 감소하였다. 5.5 mM과 22 mM의 포도당 배양액에 1, 10, 100 nM의 인슐린과 1, 10, 100 nM의 angiotensin II를 첨가한 후 대동맥평활근세포의 성장속도를 비교한 결과, 5.5 mM보다 22 mM의 포도당이 든 배양액에서 대동맥평활근세포의 성장이 촉진되었으며, 인슐린 및 angiotensin II를 첨가한 경우도 대동맥평활근세포의 성장이 증가되었다. 결론:흰쥐 대동맥평활근세포에서 PAI-1 mRNA의 발현은 포도당 농도가 높을수록 증가되며 angiotensin II의 농도 및 배양시간에 따라 증가되고 인슐린 투여로 감소하였다. 또한 angiotensin II의 투여는 22 mM의 고농도 포도당 투여 후 증가된 PAI-1 mRNA 발현 증가를 더욱 증가시켜 PAI-1 mRNA 발현 증가에 상승작용이 있음을 알 수 있다. 그리고 22 mM의 고농도 포도당, 인슐린 및 angiotensin II는 흰쥐의 대동맥평활근세포의 성장을 촉진시켰다.

• Animal cells and systems
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• 제7권3호
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• pp.249-253
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• 2003

Abnormalities in fibrinolysis system is associated with risk of hypertension. In this report, the Alu repeat insertion/deletion (I/D) polymorphism of tissue plasminogen activator (t-PA) and the Hind III RFLP of plasminogen activator inhibitor-1 (PAI-1) genes were investigated in 115 normotensives and 83 patients with hypertension, and their association with anthropometrical data and plasma biochemical parameters were analyzed. There were no significant differences in the gene frequencies of the two candidate genes between normotensives and hypertensives, respectively. Our results indicate lack of associations between the two polymorph isms in t-PA and PAI-1 genes and risk of hypertension in the population under study. However, the Hind III RFLP of PAI-1 gene was significantly associated with plasma glucose level, suggesting its role in glucose metabolism. It needs to be tested whether this RFLP of PAI-1 gene is associated with insulin resistance syndrome or non-insulin dependent diabetes mellitus (NIDDM) in the Korean population.

• BMB Reports
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• 제44권1호
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• pp.34-39
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• 2011

Resistance to PAI-1 is a factor which confers clinical benefits in thrombolytic therapy. The only US FDA approved PAI-1 resistant drug is Tenecteplase$^{(R)}$. Deletion variants of t-PA have the advantage of fewer disulfide bonds in addition to higher plasma half lives. A new variant was developed by deletion of the first three domains in t-PA in addition to substitution of KHRR 128-131 amino acids with AAAA in truncated t-PA. The specific activity of this new variant, $570\;IU/{\mu}g$, was found to be similar to those found in full length t-PA (Alteplase$^{(R)}$), $580\;IU/{\mu}g$. A 65% and 85% residual activity after inhibition by rPAI-1 was observed for full length and truncated-mutant form, respectively. This new variant as the first PAI-1 resistant truncated t-PA may offer more advantages in clinical conditions in which high PAI-1 levels makes the thrombolytic system prone to re-occlusion.

• 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.

• Tuberculosis and Respiratory Diseases
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• 제44권3호
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• pp.516-524
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• 1997

목 적 : 악성 종양에 있어서 단백질 분해 효소에 의한 세포외기질의 분해는 종양 침습 및 전이에 있어서 중요한 역할을 한다. Urokinase-type plasminogen activator(u-PA)는 이러한 단백질 분해효소 중 하나이며, u-PA의 길항제인 plasminogen activator inhibitor(PAI-1, PAI-2)도 u-PA에 의한 종양조직 자체의 분해를 방어하여 종양의 성장, 침습 및 혈관신생을 촉진하는 역할을 한다. u-PA와 PAI-1은 폐암 등의 종양 조직내의 양이 증가하며, 침습도 및 예후 둥과 유의하게 상관되는 것으로 보고되고 있다. 저자 등은 폐암환자의 혈장 u-PA와 PAI-1의 농도를 측정하여 조직형 및 병기와의 관련성을 조사해 보고자 한다. 방 법 : 폐암 환자 37예, 여러 가지 양성 폐질환 환자 21예 및 유사연령의 정상 대조군 24예에서 혈장 u-PA 및 PAI-1 항원의 농도를 ELISA법으로 측정하였다. 결 과 : 혈장 u-PA 항원 농도는 정상 대조군에서 $1.0{\pm}0.3ng/mL$, 양성 폐질환군에서 $1.0{\pm}0.3ng/mL$, 폐암환자군에서 $0.9{\pm}0.3ng/mL$로 유의한 차이가 없었다. 혈장 PAI-1 항원 농도는 정상 대조군에서 $14.2{\pm}6.7ng/mL$, 양성 폐질환군에서 $14.9{\pm}6.3ng/mL$, 폐암 환자군에서 $22.1{\pm}9.8ng/mL$로 폐암 환자에서 정상 대조군 및 양성 폐질환군에 비하여 유의하게 높았다. 혈장 u-PA 항원 농도는 편평상피암에서 $0.7{\pm}0.4ng/mL$, 선암에서 $0.8{\pm}0.3ng/mL$, 대세포암에서 0.9ng/mL, 소세포암에서 $1.1{\pm}0.7ng/mL$로 유의한 차이가 없었다. 혈장 PAI-1 항원 농도는 편평상피암에서 $22.3{\pm}7.2ng/mL$, 선암에서 $22.6{\pm}9.9ng/mL$, 대세포암에서 42ng/mL, 소세포암에서 $16.0{\pm}14.2ng/mL$로 유의한 차이가 없었다. u-PA 항원 농도는 stage I에서 0.74ng/mL, stage II에서 $1.2{\pm}0.6ng/mL$, stage IIIA에서 $0.7{\pm}0.4ng/mL$, stage IIIB에서 $0.7{\pm}0.4ng/mL$, stage IV에서 $0.7{\pm}0.3ng/mL$였고, 혈장 PAI-1 항원 농도는 stage I에서 21.8ng/mL, stage II에서 $22.7{\pm}8.7ng/mL$, stage IIIA에서 $18.4{\pm}4.9ng/mL$, stage IIIB에서 $25.3{\pm}9.0ng/mL$, stage IV에서 $21.5{\pm}10.8ng/mL$로 유의한 차이가 없었다. 폐암 환자에서 T기를 T1-3 와 T4로 나누어서 비교를 한 결과, 혈장 u-PA 항원 농도는 T1-3에서 $0.8{\pm}0.4ng/mL$였고 T4에서 $0.7{\pm}0.4ng/mL$로 차이가 없었으나, 혈장 PAI-1 항원 농도는 T1-3에서 $17.9{\pm}5.6ng/mL$였고 T4에서 $26.1{\pm}9.1ng/mL$로 T4에서 유의하게 높았다. 혈장 u-PA 항원 농도는 M0에서 $0.8{\pm}0.4ng/mL$, M1에서 $0.7{\pm}0.3ng/mL$였고, 혈장 PAI-1 항원 농도는 M0에서 $23.6{\pm}8.3ng/mL$, M1에서 $21.5{\pm}10.8ng/mL$로 유의한 차이가 없었다. 결 론 : 이상의 결과에서 폐암 환자에서 혈장 PAI-1은 정상대조군 및 양성 폐질환에 비하여 특이적으로 증가하였고, 또한 폐암의 국소적 침습 정도와의 관련성을 보여 주었다.

• Nutritional Sciences
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• 제6권2호
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• pp.73-77
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• 2003

PAI-1 (plasminogen activator inhibitor-1), leptin, and resistin are synthesized and secreted by Int cells of rodents and have recently been postulated to be an important link to obesity. This study was conducted to identify the nutritional regulation of PAI-1, leptin, and resistin gene expression in 0b/ob mice. The mice were divided into four groups according to nutritional status: control, 48 hour fasting, 48 hour-fasting/12 hour-refeeding, and 48 hour-fasting/24 hour-refeeding. The mRNA levels of each peptide were measured by semi-quantitative RT-PCR. In visceral fat tissue, the level of PAI-1 mRNA increased markedly when 48h-fasted animals were refed with a high carbohydrate-low fat diet. However, lasting/refeeding did not appreciably change PAI-1 mRNA levels in subcutaneous fat tissue. Similar results were obtained for resistin mRNA levels in both types of fat tissues. These findings suggest that visceral adipose tissue might be more sensitively involved in the nutritional regulation of PAI-1 and resistin gene expression compared to subcutaneous fat tissue. The level of leptin mRNA decreased markedly in the 48h-fasted animals, and increased markedly when 48h-fasted animals were refed with a high carbohydrate-low fat diet. The nutritional regulation of leptin mRNA showed similar patterns in both types of fat tissues. In conclusion, the nutritional regulation of gene expression encoding PAI-1, resistin, and leptin from adipocytes may vary according to the type of adipose tissue.