| IntroductionPlasminogen, a key proenzyme of plasmin, circulates as asingle-chain glycoprotein in the blood and maintains the fibrinolyticsystem. It is converted to plasmin, an active two-chain form, by tissueplasminogen activator (t-PA) in the presence of a fibrin clot orurokinase. The converted plasmin then digests the insoluble fibrinclot into soluble fragments during tissue repair and recanalization.Congenital plasminogen deficiency has been frequently reportedto be associated with thrombosis, since the reduction of plasminogenmay result in a thrombotic tendency by allowing the growth anddevelopment of thrombi. This disorder is further classified into twomajor subtypes on the basis of plasma antigen and activity levels ofplasminogen. In Type I deficiency (hypoplasminogenaemia), the levelof immunoreactive plasminogen is reduced in parallel with itsfunctional activity, whereas in Type II deficiency(dysplasminogenaemia), the level of immunoreactive plasminogen isnormal or only slightly reduced. However, there have been conflictingreports as to whether this deficiency is a significant, independent riskfactor for thrombosis.Ala601Thr mutation is thought to be relatively prevalent fordysplasminogenaemia in many types of point mutation in theabnormal genes for plasminogen. Ala601Thr mutation, type Imutation, a guanosine in GCT coding for Ala601 near the active-sitehistidine603 was replaced by an adenosine resulting in ACT coding forthreonine. Dysplasminogenemia, caused by the Ala601Thr mutationhas been found in 3.6% of the Japanese population and in 2.9% of theChinese Han population.We study the relationships between the Ala601Thr mutation andthe thrombosis by the gene frequency in DVT , CI and controls.Materials and MethodsCitrated venous blood was drawn from 120 normal individualsas well as 66 patients with deep vein thrombosis (diagnosis byultrasonography) ,67 patients with Cerebral Infarction (diagnosis byCT/MRI). Genomic DNA was amplified by 1 set of primer pairsflanking all 19 exons including intron boundaries of the humanplasminogen (sense-primer 5`ATG GAG CAG AAC AAA GTATC 3` antisense-primer 5`GAA AGG AGG AAA AGA AGGAC3`) and the PCR profiles is 94 ℃ 4min;94 ℃ 1min;59 ℃ 1min;72 ℃ 2min(2个cycle);94 ℃ 30secend;59 ℃ 30secend;72 ℃ 1min;(27个cycle);94 ℃ 1min;59 ℃ 30secend;72 ℃ 10min;4℃。For restriction fragment length polymorphism(RFLP) analysis, thePCR products were digested with Fnu4H I at 37 ℃ for 12h, separatedon 3% agarose and visualized with ethidium bromide(EB).Results and DiscussionWe detected 14 cases with heterozygous Ala601Thr mutations(10 of controls, 2 of DVT and CI respectively) and no homozygouscases by RFLP analysis.The genotypes and the allele frequency of each group has thecommunity representation carried on the Hardy-Weinbargbalance(HWE) examines(P=0.63,P=0.90,P=0.90 in cases of controls,DVT, CI respectively). The frequency of the genotype and the allelehave no statistics difference by Fisher's Exact Test to examine therelationship of the control group and DVT group, the control groupand CI group respectively(the P value is 0.217,0.225,0.218,0.226respectively).Although venous thrombosis often appear to be spontaneous inorigin, it is associated with one or more predisposing factors in thegreat majority of instances. Type II plasminogen deficiency ordysplasminogenaemia was first reported in 1978. They identified apatient with thrombosis recurring over the previous 15 years, whopossessed an abnormally low plasminogen activity but who hadnormal levels of immunoreactive plasminogen. Further studies of thepatient's family suggested that the molecular abnormality wasinherited on an autosomal gene. In 1982, they clarified that thereplacement of Ala601 by Thr in the active site of plasminogen(serine protease portion) caused a decrease of plasminogen activity.Plasminogen deficiency has been reported as a risk factor forthrombosis. In fact, recurrent thrombosis was found in the first patientwith this disorder and several publications clearly an associationbetween plasminogen deficiency and thrombosis. The observationthat plasminogen knockout mice might have a predisposition tomultiple thrombosis may be applicable to humans. In this regardindicating that heterozygotes of a Type I plasminogen deficiencywere found to experience significantly more thrombotic events(although rather late in life) than their normal family members in aretrospective analysis of 20 families. However, other reports do notsupport such an association. These reports suggest that the thrombotictendency resulting from plasminogen deficiency is still controversial.On the other hand, it has been reported that the thrombotic tendencymight be accelerated in patients with this deficiency by the presenceof additional triggering risk factors, such as factor V Leiden mutationand elevated lipoprotein(a). Although it still remains unclear whetherthe combination of this deficiency and factor V Leiden significantlyincreases the risk of venous thrombosis, it is possible that the risk ofthrombosis can be accelerated by other additional risk factors inpatients with this deficiency.Multi-gene disease is not by the simple Mendel heredity, but isthe result which affects together with the environment factor. Thethrombosis mechanism is the multi-factors, the multi-systemsparticipated together. Furthermore there are too many factor types inthe coagulation system, the anti-coagulation system and thefibrinolytic system. Therefore,it is very difficultly to appreciate theaffects that a single factor is the sick for thrombosis at present. It isthe key to explain the heredity of the thrombosis by study theconnection research of many candidate genes SNP. We experimentalresult has confirmed the multi-genes heredity special characteristicsimilarly which the thrombus forms, and provides the foundation forthe following multi-genes connection research.ConclusionThe Ala601Thr mutation is not the independence risk factor forthrombosis,The thrombosis is the result of the multi-genes heredityand the external environment affect together.
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