Jiřina Zavřelová

Clinical relevance of uPA, uPAR, PAI 1 and PAI 2 tissue expression and plasma PAI 1 level in colorectal carcinoma patients.

BACKGROUND/AIMS Urokinase (uPA) is a serine protease, which together with uPAR, tPA, PAI 1 and PAI 2 forms the plasminogen activator system, a component of metastatic cascade contributing to the invasive growth and angiogenesis of malignant tumours. METHODOLOGY Both preceding therapy and after 6-8 weeks of the treatment, plasma PAI 1 levels (photometric microplate method on the ELISA) and uPA, uPAR, PAI 1 and PAI 2 tissue expression (immunohistochemical reaction) were analysed from 80 colorectal carcinoma patients. RESULTS Analysis showed higher pre-treatment plasma levels of PAI 1 in patients with advanced tumours, which decreased after surgery or the start of therapy (p=0.004); Patients with higher plasma level PAI 1 before (0.013) and after therapy (0.004) had significantly shorter survival. There was a higher expression of uPA (p<0.001), uPAR (p<0.001), PAI 1 (p=0.042) and PAI 2 (p<0.001) in advanced colorectal carcinoma. A relationship between PAI 2 (p=0.010) and uPAR (p=0.019) expression and survival was demonstrated. There is a correlation between pre-treatment plasma PAI 1 levels and PAI 2 (p=0.028) and uPAR (p=0.043) expression. CONCLUSIONS Immunohistochemical analysis of PAS in tumour tissue and plasma PAI 1 levels was found to be a useful prognostic factor in colorectal carcinoma patients. Plasma PAI 1 could be advantageous in evaluating the effectiveness of a mode of treatment.

Increasing procoagulant activity of circulating microparticles in patients with Philadelphia-negative myeloproliferative neoplasms: a single-centre experience.

Microparticles are small membrane fragments with dimension between 0.1 and 1 &mgr;m, which are released during cell activation or apoptosis, exposing the phospholipid phosphatidylserine and membrane antigens typical for cellular origin. Philadelphia-negative myeloproliferative neoplasms (MPNs) are characterized by an increased risk of thrombosis. Data from literature suggest an association between thrombosis and the procoagulant activity of microparticles. Association between the procoagulant activity of microparticles and the incidence of thrombosis was assesed in a group of 126 patients with Philadelphia-negative MPNs. Measurement of microparticles procoagulant activity was performed using a functional assay, namely the Zymuphen MP-activity (Hyphen Biomed, Neuville-sur-oise, France). A total of 539 samples were analysed within this group of patients, regardless of patients’ state of health. A significantly higher circulating microparticles procoagulant activity was found in MPN patients as compared with the control group (P < 0.001). A pathological level of procoagulant activity was observed more frequently in patients with polycythaemia vera (88%, P = 0.002) than groups of patients with essential thrombocythaemia (73.2%) and primary myelofibrosis (68.3%); the same result was confirmed in patients with a history of venous thrombosis in comparison with patients without thrombosis (84.7 vs. 73.2%, P = 0.029). Patients without cytoreductive treatment had a higher activity of microparticles (P = 0.010). Furthermore, presence of JAK2 V617F mutation was associated with an increased procoagulant activity (P = 0.007), as well as the higher JAK2 V617F allele burden (P = 0.001). Further prospective clinical studies will be necessary to evaluate the clinical relevance of microparticles in the prediction hypercoagulable state in these patients.

A novel germline mutation in GP1BA gene N-terminal domain in monoallelic Bernard-Soulier syndrome

Abstract Mutations in the GP1BA gene have been associated with platelet-type von Willebrand disease and Bernard-Soulier syndrome. Here, we report a novel GP1BA mutation in a family with autosomal dominant macrothrombocytopenia and mild bleeding. We performed analyses of seven family members. Using whole-exome sequencing of germline DNA samples, we identified a heterozygous single-nucleotide change in GP1BA (exone2:c.176T>G), encoding a p.Leu59Arg substitution in the N-terminal domain, segregating with macrothrombocytopenia. This variant has not been previously reported. We also analysed the structure of the detected sequence variant in silico. In particular, we used the crystal structure of the human platelet receptor GP Ibα N-terminal domain. Replacement of aliphatic amino-acid Leu 59 with charged, polar and larger arginine probably disrupts the protein structure. An autosomal dominant mode of inheritance, a family history of mild bleeding episodes, aggregation pattern in affected individuals together with evidence of mutation occurring in part of the GP1BA gene encoding the leucine-rich repeat region suggest a novel variant causing monoallelic Bernard-Soulier syndrome.