A Case Of Deficiency Of Plasma Plasminogen Activator Inhibitor 1, Its Diagnosis, Treatment And Study On Molecular Abnormality

Background: Plasminogen activator inhibitor 1 (PAI-1) is the primary physiological inhibitor of plasminogen activation in blood, it is the only inhibitor found to complex with single-chain tissue type-plasminogen activator (tPA). Delayed bleeding occurs because a normal hemostatic plug is formed, but it is later more susceptible to breakdown owing to increased fibrinolytic activity. Several PAI-1-deficient patients with decreased or undetectable PAI-1 activity or antigen have been reported in literature. In this report, we present a patient with low plasma levels of PAI-1 activity and antigen, which was associated with a lifelong delayed bleeding disorder.Patient: The patient, male, born in 1970, had recurrent episodes of subcutaneous bleeding starting in early childhood, all in response to trauma or surgery. No complications were reported at birth. At the age of sixteen years, he had left leg minor trauma that resulted in a subcutaneous hematoma, then muscular atrophy. At thirty years, a big subcutaneous hematoma caused his right leg stiflhess. An operation was performed in order to clear the organized clot, but delayed bleeding after surgery lasted about four months, and required several tunes of fresh plasma transfusion, the amount being up to 2800 ml. One month before his hospitalization, a trauma resulted in two hematomas in his left leg and left hip. There was no history of spontaneous hemorrhage or hemarthrosis, and the family history was negative for abnormal bleeding. He had numerous hemostatic evaluations but no definite diagnosis. The patient was physically and developmentally normal. Physical examination of the patient showed no particular abnormalities.Hemostatic tests: Hemostatic screening test results were normal. Bleeding time was 7min, activity of factor VIII ,factor IX, and factor XIII was 72.9%, 96.1% and 100%, respectively. D-dimer and fibrin degradation products (FDP) were within normal range. The patient had normal platelet counts and morphology. Platelet aggregationresponses to agents (ADP, thrombin, collagen, epinephrine, and ristocetin) were within normal range. Euglobulin clot lysis time (ECLT) was 70 minutes, shorter than normal. PAI-1, tPA and a 2-antiplasmin ( a 2-AP) activity were measured by chromogenic assay, PAI-1 and tPA antigens were measured by enzyme-linked immunosorbent assay (ELISA). Activity of a 2-AP was within normal range. Concentrations of tPA activity and antigen in patient's plasma were 0.33AU/ml and 20.4ng/ml, respectively, within normal range. Concentrations of PAI-1 activity and antigen were 0.04 AU/ml and 5.6ng/ml, which dramatically decreased (normal control values were 0.41 AU/ml and 44ng/ml, respectively). After the diagnosis of a PAI-1 deficiency was established, the patient was treated with aminomethylbenoic acid (0.4g~0.6g, q6h ?q8h) before and after operation, surgery was performed successfully with no excessive blood loss, plasma and red cell concentrate were not given during surgical procedure, delayed bleeding was significantly alleviated than before.Study on genetic abnormality: To clarify the cause of the genetic abnormality responsible for PAI-1 deficiency phenotype in the patient, the genetic DNA of the patient was extracted, the genes encoding PAI-1 exon were amplified by polymerase chain reaction (PCR), and direct nucleotide sequencing was carried out on PCR products using a DNA sequencing kit. PCR products were cloned in pUCm-T vector, and transformed into E. Coli TGI competent cells, the white clones were sequenced in both directions. The results were blasted with GenBank, nucleotide sequencing revealed that the patient had heterozygous missense mutation in exon 2, a G to A transition at nucleotide 43, causing AlalSThr at signal peptide. The clone sequencing showed the patient had both G and A at that site. The restriction endonuclease (RE) analysing showed that heterozygosity were found in patient and his father, but not in 60 cases of normal subjects and his other family members. This excluded the possibility that the mutation was a pol...