Search published articles
Showing 3 results for Tissue Plasminogen Activator
- -, - -, - -,
Volume 5, Issue 2 (8-2014)
Abstract
Tissue plasminogen activator (tPA) is a 65 kDa member of serine protease family. tPA is naturally expressed by endothelial cells of vessels in negligible amounts. The enzyme converts plasminogen to plasmin and results in dissolution of blood clots. Retepalse (Retavase) is a mutated variant of human tPA that lacks a segment of 137 residues. Reteplase is produced in Escherichia coli and lacks post translational modifications including glycosylation. Due to presence of nine disulfide bonds, expression of this protein within bacterial systems is so difficult and mostly results in protein aggregates. Refolding and reactivation of inclusion bodies is an expensive, time-consuming procedure with low efficacy for disulfide-bonded proteins. In the present research we have changed the regulatory sequences to produce active and soluble reteplase enzyme in E. coli BL21 host. Inducible expression of reteplase under the control of T7 promoter resulted in aggregation of proteins in inclusion bodies while the use of a constitutive promoter could produce biologically active reteplase in the cytosol of E. coli cells.
Volume 16, Issue 1 (8-2013)
Abstract
Objective: Development of high producing mammalian cell lines is a major bottleneck in manufacturing of recombinant therapeutic proteins. This study examines the effect of using the matrix attachment region from the human interferon beta gene in combination with promoter activation strategy with E1A 13S protein on human tissue plasminogen activator (t-PA) expression in Chinese hamster ovary (CHO) cells. Methods: The matrix attachment region was cloned in 3΄ and 5΄ flanking sides of the t-PA expression cassette in pTPA vector to generate pMTPA. After transfection of the cells with pTPA and pMTPA vectors, stable cell pools were developed and the t-PA expression level determined for each stable cell line. In the next step, E1A 13S expression plasmid was transfected to stable cell pools and t-PA titers were measured after 72 hours. Results: Integration of pTPA and pMTPA vectors in the CHO genome was confirmed by PCR analysis on genomic DNA of stable cell pools. Analysis of the t-PA expression level showed a three-fold enhancement in pMTPA transfected cells compared to pTPA-containing cells. t-PA expression was further enhanced up to 1771 U/ml by transient expression of E1A 13S in pMTPA stable cell pools. Conclusion: These results have shown that incorporation of matrix attachment region in an expression vector in combination with promoter activation can effectively enhance recombinant protein expression levels in CHO cells.
Volume 21, Issue 3 (10-2018)
Abstract
Aims: Tissue factor pathway inhibitor (TFPI) and tissue plasminogen activator (tPA) deficiency would cause an increase in the viscosity of fibrinogen and low digestion of blood clot, respectively, which are among the dangerous factors responsible for the increase in thrombosis. Although there are reports, indicating the relationship of vein thrombosis and recurrent pregnancy loss (RPL), the effect of polymorphisms connected with thrombosis on the risk of increasing RPL has remained unclear. Thus, the aim of this study was to evaluate the relationship of 536C/T tissue factor pathway inhibitor gene and Alu_I/D tissue-type plasminogen activator gene polymorphisms with recurrent pregnancy loss in Iranian Azeri women.
Materials and Methods: The present case-control study was conducted in Azari women with RPL and healthy women during 2015-2016. Ninety-five women with RPL were selected by simple random sampling sampling method, and 95 healthy women were selected and assessed, using Multiplex ARMS-PCR method. The data were analyzed by SAS 9.2 software, using logistic regression and Chi-square test.
Findings: Among two genes polymorphisms, only tPA polymorphism was significantly related with the increase in RPL, such that allele I in this polymorphism caused more than two-folded odds ratio of RPL compared to allele D. Women between the ages of 25 and 35 years significantly showed the highest odds ratio of RPL.
Conclusion: Alu_I/D tissue-type plasminogen activator gene polymorphism is related with an increased risk of RPL in Azeri women in northwestern Iran, but 536C/T tissue factor pathway inhibitor gene does not show such a relationship.
