Analysis The Human Platelet Alloantigen Frequency Of Hebei Han Population

Objective: Human platelet-specific alloantigen(HPA) carried by platelet glycoprotein, located mainly on the membrane of platelets, is gene single nucleotide polymorphism. As the widespread use of the platelet products in the clinical application, platelet alloantibodies, leading alloimmune thrombocytopenia, such as blood transfusion after induced thrombocytopenic purpura(PTP), platelet transfusion refractoriness(PTR) and newborn children alloimmune thrombocytopenic purpura(NAITP) and other diseases occurred because of platelet-specific antigen mismatched platelets and the same kind of immune response. Meanwhile in clinical transplantation also cause graft rejection and other related diseases. HPA antibodies are also the main cause of the disease such as neonatal alloimmune thrombocytopenic purpura(NAITP) and post-transfusion thrombocytopenic purpura(PTP) disease. For some random platelet transfusions in patients with platelet transfusion produced invalid, HLA antigens compatible platelet transfusion also failed to improve the condition; In some patients platelet transfusion refractoriness, no antibody was detected cytotoxicity, these two points are suggestive of anti-platelet antibodies in these patients in vivo, Therefore, accurate detection and identification of HPA, and for clinical transfusion practice is quite important.Population genetics studies show that the distribution of HPA gene has obvious racial and geographical differences. With blood components used in the clinical, HPA gene frequency surveys for clinical transfusion practice has important significance to help solve the problem of ineffective platelet transfusion, and can be distributed in different regions according to predict gene frequencies among different ethnic platelet-specific the probability of occurrence of immune isotype. Currently, the genetic polymorphism of HPA system on Chinese population is rarely reported, and there are no large sample volumes on the Han population in Hebei accurate genotyping data.The purpose of this study is to analysis the distribution of polymorphisms in Han population in Hebei platelet donor genes, to explore Hebei HPA gene frequencies in Han population, to establish localized platelet donors database for patients, which will provide compatible platelets, avoide allo-antibodies, and prevent from occurrence of platelet transfusion refractoriness.Methods:Preparation of genomic DNA: samples were from a random sample of 930 unrelated Hebei Han healthy platelets donors. 5ml anticoagulated blood was taken with EDTA tube. The genomic DNA was prepared by the TIANGEN blood DNA purification kit.1 The extracted DNA should meet OD260 / OD280 ratio between 1.7-1.9, the concentration of 30 ng /μl or so.2 Allele typing: Using the HPA-1-17 genotyping PCR kit of Tianjin Super Biotechnology Developing Co.,Ltd, according to the HPA alleles provided by NCBI Gene Bank, using PCR-SSP, HPA1-17 alleles genotype were analyzed.3 PCR product was analyzed by gel electrophoresis: The PCR products were transferred to agarose gel electrophoresis, the results observed in the UV /gel imaging system and shooting imaging, according to the results of interpretation of genotyping primer hole provided in the bitmap.4 Statistical analysis: gene using gene frequency is calculated by direct counting method, and Hardy-Weinberg(HW) test to verify the goodness of fit. The mismatch rate of each system HPA dual antigen using the formula P = 2ab(1-ab) calculations(a and b represent the gene frequency). The gene frequency of 930 Han population in Hebei were compared with other parts of the Han population and domestic and foreign populations in other regions by χ2 testResults:1 930 HPA genotyping results in Hebei Han platelet donors are as follows: 919 for HPA-1aa, 11 for HPA-1ab, 0 for HPA-1bb; 861 for HPA-2aa, 69 for HPA-2ab, 0 for HPA-2bb; 339 for HPA-3aa, 421 for HPA-3ab, 170 for HPA-3bb; 928 for HPA-4aa, 2 for HPA-4ab, 0 for HPA-4bb; 911 for HPA-5aa, 19 for HPA-5ab, 0 for HPA-5bb; 919 for HPA-6aa, 10 for HPA-6ab, 0 for HPA-6bb; 294 for HPA-15 aa, 358 for HPA-15 ab, 278 for HPA-15bb; on the HPA-7,8,9,10,11,12,13,14,16,17 sites are all aa homozygous genotype among the 930 donors and b gene were not detected.2 HPA gene frequencies obtained from 930 in Hebei Han platelet donors are as follows: 99.41% and 0.59 for HPA-1a and HPA-1b; 96.29% and 3.71% for HPA-2a and HPA-2b;59.09% and 40.91% for HPA-3a and HPA-3b;99.89% and 0.11% for HPA-4a and HPA-4b; 98.98% and 1.02% for HPA-5a and HPA-5b; 99.46% and 0.54 for HPA-6a and HPA-6b; 50.86% and 49.14% for HPA-15 a and HPA-15b; HPA-7a, 8a, 9a, 10 a, 11 a, 12 a, 13 a, 14 a, 16 a, 17 a gene frequency was 100%, which corresponds to a frequency of 0 b gene.Conclusions:1 After a large number of samples of experimental manipulations, PCR-SSP method was proved the preferred method for the detection of antigen HPA conducted.2 The population distribution of HPA allele has a significant difference in ethnics and geography, and the HPA gene has a very rich polymorphism in Hebei Han population. Higher rates do not match with the Han population in Hebei HPA-3, 15, and the rates are more than 37%. Additionally, opportunity to generate anti-platelet amtibody was increased in the platelet transfusion, suggesting that HPA-3, 15 is the main element to induce Hebei Han platelet transfusion reactions, which should pay more attention in platelet transfusion.3 HPA-1 gene frequency of Hebei Han population had a significant difference compared to that of Europe, African, although HPA- 1, 3, 15 gene frequencies of Hebei Han population had no statistically different from most regions of the our country. In particular, HPA- 2, 5 gene frequencies of Hebei Han population were statistically different from that of other regions at home and abroad, indicating that gene distribution in Hebei HPA has its own characteristics4 Hebei Han population HPA gene frequency data in this study has taken an important step for the construction of the Hebei region investigated platelet donor bank, and laid the foundation for solving platelet transfusion, as well as provided reference materials for anthropology and population genetics in the region research and other aspects.