Study on the extensibility of platelet donor gene database in Shaanxi / 中国输血杂志

【Objective】 To analyze the commonality and characteristics between voluntary blood donors and hematopoietic stem cell donors in this region, and explore the potential for integration and development between China Marrow Donors Program (CMDP) and voluntary blood donors, especially platelet donor databases, so as to improve recruitment success rate and inventory rate. 【Methods】 The database modeling and comparison methods were used to screen and stratify the matching and integration degree between the voluntary blood donors in recent 10 years and the marrow donors in the Shaanxi Branch of CMDP. The frequencies of HLA-A,-B alleles, HPA alleles and haplotypes were calculated with Arlequin 3. 5. 2. 2 software, and the matching probability of different platelet donor reserve pools was conducted according to the phenotypic frequencies. 【Results】 Among the voluntary donors with known HLA genotypes in this region, according to their blood donation behavior,the active blood donors excavated were divided into the first, second, third and fourth echelons of platelet donor reserve pools, with 696, 2 752, 9 092 and 12 028 donors, respectively. The first echelon had the highest proportion of 10-50 times of platelet donations and 10-20 times of whole blood donations, with 13.65% and 26.01%, respectively. The second echelon had 10-20 times of whole blood donations and 10-50 times of platelet donations, accounted for 15.04% and 1.38%, respectively, which were significantly different from other echelons' blood donation characteristics (P<0.05). With a database size of the existing platelet donor bank adding the first and second echelons (n=4 955), there was a 69.02% probability of matching at least one donor with matching HLA-A-B phenotype. When considering the matching ABO and HPA phenotypes, the probability of finding at least one donor with fully matching HLA, HPA and ABO isotype (type B as an example) was 48. 73%. 【Conclusion】 The three groups of whole blood donation, apheresis platelet donation and marrow donation in Xi'an area have a large cross-distribution. Compared with expanding the storage capacity from scratch, the active blood donors in CMDP database are the largest back-up force of platelet donors. While expanding the effective storage capacity, it can minimize the cost of building platelet donor bank and the demand for resources.

The polymorphism of HPA-1-6w, HPA-15 and HPA-32bw-35bw in platelet donors from Deyang, Sichuan / 中国输血杂志

【Objective】 To explore the polymorphism of HPA-1-6w, HPA-15 and 32bw-35bw in platelet donors in Deyang, Sichuan, and estimate whether to include the detection of 32bw-35bw in the platelet bank. 【Methods】 Polymerase chain reaction with sequenced based typing (PCR-SBT) was used to sequence the HPA-1-6w, HPA-15 and 32bw-35bw loci of 205 platelet donors in Deyang. Allele frequencies were calculated by the direct counting method. The frequencies of HPA-1-6 and 15 alleles in northern and southern Chinese, Japanese and Australian population were compared, and those HPA loci and HPA-32bw-35bw were searched in the Chinese Millionome Database (CMDB) and genomAD to obtain the polymorphism data. Then the Chi-square test was performed with the data of this study through GraphPad Prism 9 software. 【Results】 The allele frequencies of HPA-1b, 2b, 3b, 5b, 6bw and HPA-15b were 0.005(2/410), 0.037(15/410), 0.471(193/410), 0.020(8/410), 0.010(4/410) and 0.461(189/410), respectively, b allele of HPA-32bw-35bw and HPA-4 was not detected. Statistical significance was observed between the HPA-1b allele frequency of this study and northern Chinese, Australian population and genomAD global population sample (P< 0.05, 0.005 vs 0.014 vs 0.145 vs 0.122). The frequency of HPA-2b alleles in this study, Japanese population and genomAD global population samples was 0.037 vs 0.120 vs 0.100, with statistical difference(P<0.05). Comparison of HPA-5b and HPA-6bw allele frequencies with those of genomAD global population showed a statistical difference (P<0.05, 0.020 vs 0.089 and 0.010 vs 0.000 008, respectively). 【Conclusion】 The polymorphisms of HPA-1-6w and HPA-15 of donors in Deyang has characteristics of the southern Chinese. The frequencies of HPA-32bw-35bw were extremely low, which could be excluded from the platelet bank in Deyang.

The capacity evaluation and utilization strategy of the platelet donor database with known HLA/HPA genotype in Shaanxi / 中国输血杂志

【Objective】 To evaluate the appropriate optimal capacity and matching probability of the platelet donor database with known HLA/HPA genotype in Shaanxi aera, and provide data support for subsequent construction, maintenance and application of the local platelet donor database. 【Methods】 A total of 11 755 individuals from the Shaanxi Branch of China Marrow Donor Program, 401 and 249 unrelated random platelet donors in Shaanxi aera were enrolled to the population study of HLA-A, -B polymorphisms, HPA genotyping and CD36 antigen expression, respectively. The frequencies of HLA-A, -B alleles, HPA alleles and haplotypes were calculated with Arlequin 3. 5. 2. 2 software; matching probability and capacity evaluation of platelet donor database was conducted according to the phenotypic frequencies. 【Results】 The population genetic and phenotypic polymorphisms data of HLA-A, -B and HPA1-6, 10, 15, 21 in Shaanxi aera were obtained. The frequency of CD36 type Ⅰ or Ⅱ deficiency was 0.40%(1/249). According to the subsequent calculating and deriving, with a database size of 194 donors, the patient having approximate 95% probability could achieve matching of HPA1-6, 10, 15, 21 genotype. With a database size of 1500 donors, there is a 95% probability of matching at least one donor with HLA-A-B phenotype frequency >0.002 or haplotype frequency >0.001; meanwhile, the probability of matching a cross-reactive group donor should be 44.95%-97.57%. Based on database size of 8 856 and 15 033, the probabilities of matching HLA-A, -B phenotype were about 80% and 90%, respectively. 【Conclusion】 The differences in the distribution of HLA/HPA polymorphism in different regions make the establishment mode and optimal capacity of platelet donor database different. It is necessary to apply a variety of platelet matching transfusion strategies to expand the range of donor selection, thereby effectively reducing the database construction cost and resource requirements.

Analysis and prospect of platelet compatible transfusion / 中国输血杂志

Platelet compatible transfusion can effectively solve the immune mediated platelet transfusion refractoriness (PTR), save platelet resources and improve blood safety. This paper comments and prospects the compatibility modes of HLA, HPA and CD36, HLA antibody titer, antigen immunogenicity and the development of platelet compatible transfusion. The pattern of HLA compatible platelets involves the matching in the alleles, antigens and epitopes levels, respectively, as well as avoidance donor specificity antibody (DSA) method. While setting the mean fluorescence intensity (MFI) threshold of avoidance DSA needs to be explored when using the DSA prediction method. Allele specific HLA antibodies can be found in the patients with PTR. Therefore, the patients and donors should be genotyped for HLA-A, -B loci at high-resolution level in order to avoid allele specific HLA antibodies. The immunogenicity of various antigens or epitopes at HLA-A and -B loci are different. Selecting donor platelets with low antigen expression or low immunogenicity may be a way of HLA compatible platelets. As the probability and type of HPA antibody production are different in the various populations, the approaching of compatibility HPA involves allele matching and avoidance DSA. As to CD36, the compatibility mode mainly refers to avoidance DSA, which means blood donors with CD36 antigen type Ⅰdeficiency are preferentially selected, and then those with CD36 antigen type Ⅱ deficiency. In the future, more attention should be paid to the scale up of database capacity and update of the information construction. The time waiting for compatible platelets transfusion in clinical could be significantly shortened if the requiring and matching are only conducted within the inventory and candidate platelets.

Allele and haplotype frequencies of human platelet and leukocyte antigens in platelet donors / Frequência alélica e haplotípica dos antígenos plaquetários e leucocitários humanos em doadores de plaquetas

ABSTRACT Objective To described the allele and haplotype frequencies of human leukocyte antigen genes at the -A, -B loci and human platelet antigen genes for human platelet antigen systems 1 to 9, 11 and 15 in blood. Methods We included 867 healthy unrelated volunteer donors who donated platelets between January 2011 and December 2014. Microarray genotyping was performed using a BeadChip microarray. Medium resolution typing of the human leukocyte antigen at loci A and B was carried out using sequence-specific oligonucleotide probe hybridization. We used multivariate analysis and our human leukocyte antigen population was compared to data from the United States national bone marrow donor program. Human platelet antigen results were compared to a literature review and data from around the world. Results Our human leukocyte antigen haplotype results were more similar to those of hispanics, followed by caucasians. Likewise, our human platelet antigen sample is more similar to those of Argentina, Rio Grande do Sul and Italy. Conclusion This was the first article that discusses human platelet antigen and human leukocyte antigen data together. Rare genotypes or antibody associations can make patient management difficult. A blood bank with genotyped donors allows for optimal transfusion and can contribute to better results. Our information can serve as basis for a database of platelet antigen polymorphisms.

RESUMO Objetivo Descrever as frequências alélicas e haplotípicas de genes dos antígenos leucocitários humanos nos loci -A,- B e dos antígenos plaquetários humanos para os sistemas HPA-1 a 9, 11 e 15. Métodos Foram incluídos 867 doadores voluntários, saudáveis, não relacionados, que doaram plaquetas por aférese entre janeiro de 2011 e dezembro de 2014. A genotipagem foi realizada usando microarray BeadChip. A tipificação de resolução intermediária dos antígenos leucocitários humanos loci A e B foi realizada por meio de hibridização com sonda para oligonucleotídeos por sequência específica. Utilizamos análises multivariadas e o antígeno leucocitário humano de nossa população foi comparado com a do programa nacional de doadores de medula óssea norte-americano. Já os resultados dos antígenos plaquetários humanos foram comparados à revisão da literatura e a dados de populações de outros países. Resultados Os resultados do haplótipo de antígenos leucocitários humanos são mais parecidos com os dos hispânicos, seguidos dos caucasianos. Igualmente, a amostra de antígenos plaquetários humanos foi mais semelhante às da Argentina, do Rio Grande do Sul e da Itália. Conclusão Este foi o primeiro artigo a discutir antígenos plaquetários e leucocitários humanos simultaneamente. Genótipos raros ou associações de anticorpos podem dificultar o manejo clínico do paciente. Um banco de sangue com doadores genotipados permite um melhor resultado e transfusão possíveis. Estas informações podem servir de base para um banco de dados sobre polimorfismos de antígenos plaquetários.

Establishment of a direct detection method for human platelet antigen1-17,Cab allele whole blood / 国际检验医学杂志

Objective To establish an acurate and convenient method to distinguish human platelet antigen(HPA) SNPs based on Target Enriched Multiplex-PCR(TEM-PCR),fluorescent probe melting curve analysis and blood direct PCR.Methods Design TEM-PCR primers and probes of HPA1-17,Cab alleles,amplify target sequences of all 18 alleles by blood direct PCR and distinguish different SNPs by melting curve of probes.Results The TEM-PCR could amplify all target sequences of 18 alleles and the melting curve analysis could distinguish those SNPs,the accuracy was equal to PCR-SSP method and the process was more convenient without blood genomic DNA extraction and subsequent gel electrophoresis thus decrease the cross-contamination risk.Conclusion Successfully established a HPA1-17,Cab alleles distinguishing method based on TEM-PCR,blood direct PCR and fluorescent probe melting curve analysis technique.

The absence of the human platelet antigen polymorphism effect on fibrosis progression in human immunodeficiency virus-1/hepatitis C virus coinfected patients

Hepatic fibrosis progression in patients with chronic hepatitis C virus infections has been associated with viral and host factors, including genetic polymorphisms. Human platelet antigen polymorphisms are associated with the rapid development of fibrosis in HCV-monoinfected patients. This study aimed to determine whether such an association exists in human immunodeficiency virus-1/hepatitis C virus-coinfected patients.

Human Platelet Antigen Genotyping Using a Multiplex Single-Base Primer Extension Reaction in Koreans / 대한수혈학회지

BACKGROUND: Alloimmunization of human platelet antigens (HPA) is associated with clinically significant disease, such as platelet refractoriness, neonatal alloimmune thrombocytopenia, or posttransfusion purpura. It is determined by single nucleotide polymorphism of genes for platelet membrane glycoprotein. To date, approximately 27 HPAs have been discovered, and their frequencies differ depending on ethnicity and country. METHODS: We conducted an investigation of prevalence of HPA in the Korean population using a multiplex single-base primer extension reaction (SNaPshot). With 84 specimens from healthy donors, HPA genotyping was performed on 11 different HPAs, including HPA-1, -2, -3, -4, -5, -6, -7, -8, -9, -13, and -15. RESULTS: A total of 90 blood samples were genotyped. The genotype frequencies of HPA were as follows: HPA-1a/1a: 100.0%, -2a/2a: 83.3%, -2a/2b: 14.3%, -2b/2b: 2.4%, -3a/3a: 39.3%, -3a/3b: 52.4%, -3b/3b: 8.3%, -4a/4a: 100.0%, -5a/5a: 95.2%, -5a/5b: 4.8%, -6a/6a: 94.0%, -6a/6b: 6.0%, -7a/7a: 100.0%, -8a/8a: 100.0%, -9a/9a: 97.6%, -9a/9b: 2.4%, -13a/13a: 100.0%, -15a/15a: 23.8%, -15a/15b: 51.2%, and -15b/15b: 25.0%. CONCLUSION: The SNaPshot assay was employed for detection of SNPs in various clinically significant HPA genes. In addition to well-known frequencies of previously reported HPA-1 to -8, this study showed frequencies of HPA-9, -13, and -15 in Koreans for the first time. The SNaPshot technique might be suitable for use in actual clinical testing in patients with platelet alloimmunization.

Study of the genetic and polymorphism of human platelet antigens 1—6,15 in unrelated volunteer blood donors,Guangzhou / 中国输血杂志

Objective To study the polymorphism of human platelet antigens among unrelated Guangzhou blood donors with PCR-SSP,in order to provide basic data for population studies and clinical transfusion practice.Methods Blood samples from 706 unrelated blood donors in Guangzhou were genotyped for each of the HPA1—6,15 systems by PCR-SSP.Gene frequencies and genotype frequencies were analyzed by statistical methods.Results HPA-3 and-15 had the greatest heterozygosity with a gene frequency of 0.2918,0.4830,0.2252 for HPA-3a/a,HPA-3a/b,HPA-3b/b,and 0.2691,0.5170,0.2139 for HPA-15a/a,HPA-15a/b,HPA-15b/b.The a/a homozygosity was predominant in HPA-1,-2,-4,-5,with a frequency ranged from 0.9583 to 0.9993,while HPA b/b was not found among them.The frequency of HPA-lb and HPA-4b was very low,which was 0.0028 and 0.0007,respectively.In our study,HPA-1 frequency was significantly different from that of the north Chinese,English,and American Indian(P

Detection of platelet specific antigens by flow cytometry and genotyping method / 대한수혈학회지

BACKGROUND: To identify the human platelet antigens (HPA) associated with neonatal alloimmune thrombocytopenia (NATP), posttransfusion purpura (PTP), and platelet refractoriness, polymerase chain reaction-sequence specific primer (PCR-SSP) method and immunofluorescent method by flow cytometry were used. The frequencies of the genonotypes of HPA systems by PCR-SSP method and those of phenotypes by flow cytometry were determined. Then both types were compared each other and each types were compared with those of established reports. METHOD: Platelet suspensions were prepared from peripheral blood specimens of 200 blood donors and DNA specimens were extracted from those of 160 donors among them. Phenotypes of 200 specimens and genotypes of 160 ones were tested by flow cytometry and PCR-SSP method, respectively. RESLUTS: Frequencies of penotypes of HPA-1a, -3a, -4a, -4b and NaKa were 100.0%, 88.0, 100.0%, 0.5% and 94.0%, respectively. HPA-5 system could not be identified due to a few antigenic sites of HPA-5 systems. The genotype fequencies are of HPA-2 were a+b- 63.75%, a+b+ 35.00%, a-b+ 1.25%; HPA-3, a+b- 38.12%, a+b+ 48.13%, a-b+ 13.75%; HPA-4, a+b- 100.00%, a+b+ 0.00%, a-b+ 0.00%; HPA-5, a+b- 98.12%, a+b+ 1.88%, a-b+ 0.00%. The frequencies of HPA-4 and -5 were almost same as those of other reports but the frequencies of HPA-2 and -3 were somewhat different from others. Concordant rate between phenotype and genotype of HPA-3a,-4a and -4b were 95.6%, 100% and 99.4%, respectively. CONCLUSION: Phenotyping method by flow cytometry was rapid and objective for identifying HPA systems except HPA-5 system which has a few antigenic sites on platelet membrane. Especially it will be useful method for screening HPA-4b as possible cause of NATP in Koreans. But like other serologic methods, phenotyping by flow cytometry also require the highly qualified antiserum and appropriate amount of platelet. PCR-SSP method was also rapid and simple to test of genotypes of HPA-2~5 systems. Because PCR-SSP method is thought to be one of the most simple and economic genotyping methods to overcome the shortages of serologic methods, it is suggested to be the efficient screening method of HPA systems substituting the serologic methods in the cases which HPA sytems can not be identified by flow cytometry.

Establishment of eight human platelet antigen systems and HLA-Ⅰ antigen donor bank in Guangzhou area / 中国输血杂志

Objective To establish eight human platelet antigen systems and HLA-Ⅰ antigen donor bank,and to determine the gene frequencies of human platelet antigen(HPA) and HLA-Ⅰin Guangzhou area.Methods A total of 805 blood samples from Chinese Han voluntary platelet donors were included in this study.PCR-SSP was used to detect single-nucleotide polymorphism in HPA systems.Luminex-SSO was used to detect the HLA-Ⅰantigens.Results The distribution of HPA 1,2,3,4,5,6,9,15 was in Hardy-Weiberg equilibrium among study subjects.Allele frequencies of 0.998 and 0.002 were observed for HPA 1a and 1b,0.952 and 0.048 for HPA 2a and 2b,0.553 and 0.447 for HPA 3a and 3b,0.999 and 0.001 for HPA 4a and 4b,0.976 and 0.024 for HPA 5a and 5b,0.982 and 0.018 for HPA 6a and 6b,1 and 0 for HPA 9a and 9b,0.518 and 0.481 for HPA 15a and 15b.The high frequency HLA-Ⅰ alleles were A*02,0.286;A*24,0.162;A*11,0.323;B*46,0.147;B*75,0.100;C*01,0.177;C*03,0.289;C*07,0.179.Conclusions This study confirmed the ethnic and territorial difference of HPA and HLA-Ⅰ.The establishment of HPA and HLA-Ⅰ matched plateletpheresis donor registry is helpful in the improvement in platelet transfusion.

Detect and characterize platelet-reactive antibodies in patients who were refractory to platelet transfusions / 中国输血杂志

Objective To detect and determine the specificity of platelet-reactive antibodies in patients who were refractory to platelet transfusions.Methods Serum samples from 48 patients who were refractory to platelet transfusions were screened with MACE for platelet-reactive antibodies.Specificity of platelet alloantibodies was determined with PAK12 and MAIPA.Results Platelet-reactive antibodies were detected in the serum of 50% PTR patients(24/48).The incidence of HLA antibodies was 39.6%(19/48),accounting for 79.2% of serum with platelete alloantibodies.The HPA alloantibodies were found in 29.2%(14/48)serum,of which,64.3%(9/14)occurred together with anti-HLA.The following platelet-specific antibodies were identified:anti-HPA-3a(n=2),anti-HPA-5b(n=1),anti-HPA-5a(n=1),anti-HPA-2b(n=1),anti-HPA-4b(n=1).Of the 14 serum with HPA antibodies,78.6%(11/14)contained panreactive anibodies against platelet glycoprotein(GP)Ⅱb/Ⅲa,GPⅠa/Ⅱa,and/or GPⅠb/Ⅸ.Platelet-reactive antibodies were detected more in female(16/29)than in male(8/19)with a frequency of 55.2%,42.1%,respectively,but there was no statistical significant difference.Conclusion The platelet-specific antibody in PTR patients are not as rare as previous thought although alloantibodies are predominantly anti-HLA.Antibody specificities in Chinese PTR patients are different from those observed in Caucasians,in whom anti-HPA-5b and-1b are the most prevalent specificity.The most prevalent platelet-specific antibodies are anti-HPA-3 and anti-HPA-5 while anti-HPA-4b and anti-HPA-2b are also detected.

Genetic polymorphism of HPA-1 to-5,HPA-15 alloantigen system in Qingdao Han population / 中国输血杂志

Objective To study the polymorphism of human platelet antigen HPA-1 to HPA-5,and HPA-15 system in Qingdao Han population.Methods A total of 918 samples from regular voluntary platelet donors in Qingdao were genotyped for HPA-1 to-5 and HPA-15 by PCR-SSP.Results The gene frequencies of HPA-1a,-1b;HPA-2a,-2b;HPA-3a,-3b;HPA-4a,-4b;HPA-5a,-5b;HPA-15a,-15b were 0.9940,0.0060;0.9319,0.0681;0.5822,0.4178;0.9897,0.0104;0.9804,0.0196;0.4913,0.5087,respectively.Both a and b alleles were found in each of the 6 HPA systems,and a/a homozygosity was more common in HPA-1,-2,-4 and-5 systems.The HPA genotype frequencies followed Hardy-Weinberg principle.HPA-1 frequency of Qingdao people was significantly different from that of North China(P