| Part 1: Genetic polymorphism analysis of clopidogrel metabolism related genes in Han and Miao populationsObjective: To analyze the polymorphism of clopidogrel metabolism related genes in the Han nationality and Miao nationality,and to explore the distribution.Methods: Han and Miao patients were collected randomly as the research object.And they were drew venous blood(2ml)for DNA extraction after signing the informed consent form.Polymerase chain reaction(polymerase chain reaction,PCR)technique was used to amplify the target DNA,and 2% agarose gel electrophoresis was used to determine whether DNA was amplified and whether the gene segment was consistent with the expected results.Then the PCR products were directly sequenced.According to the sequencing results,the gene polymorphism was analyzed.The genes include CYP2C19,ABCB1,CYP3A4,CYP3A5.Results: A total of 115 patients were collected.There were 37 males and 24 females in Han population,with an age of 49.76±16.5.And there were 32 males and 22 females in Miao population,with an age of 68.7±11.2.In Han population,the GG,GA,AA genotypes on CYP2C19*2 were: 18 cases(19.51%),30cases(49.18%),13 cases(21.31%);the GG,GA,AA genotypes on CYP2C19*3 were: 61 cases(100%),0 cases(0%),0 cases(0%);the CC,CT,TT genotypes on ABCB1(C3435T)were: 30 cases(49.18%),25 cases(40.98%),6 cases(9.84%);the GG,GA,AA genotypes on CYP3A4(IVS10+12G>A)were:30 cases(49.18%),29 cases(47.54%),2 cases(3.28%);the TT,TC,CC genotypes on CYP3A5(rs3800959): 43 cases(70.49%),16 cases(26.23%),2cases(3.28%).In Miao population,the GG,GA,AA genotypes on CYP2C19*2were: 11 cases(20.37%),33 cases(61.11%),10 cases(18.52%);the GG,GA,AA genotypes on CYP2C19*3 were: 54 cases(100%),0 cases(0%),0cases(0%);the CC,CT,TT genotypes on ABCB1(C3435T)were: 16cases(29.63%),28 cases(51.85%),10 cases(18.52%);the GG,GA,AA genotypes on CYP3A4(IVS10+12G>A)were: 21 cases(38.89%),29cases(53.70%),4 cases(7.41%);the TT,TC,CC genotypes on CYP3A5(rs3800959): 44 cases(81.48%),9 cases(16.67%),1 cases(1.85%).The frequency of ABCB1(C3435T)type CC showed statistically difference between the two ethnic groups(P=0.033),and the frequency of allele T was also showed significantly difference(P=0.027).There were no statistical differences between the two groups in terms of the other genotypes.Conclusion: The mutation frequency of CYP2C19*2,ABCB1(C3435T),CYP3A4(IVS10+12G>A)was a little high,but of CYP3A5(rs3800959)was low.And there was no mutation on CYP2C19*3.The mutation frequency of ABCB1 was significantly higher in the Miao population than in the Han population,while the frequency of CYP2C19 and CYP3A4/5 mutations were not significantly different between the two groups.However,the number of samples was small.It still need to increase the samples for further verification.Part 2: Relationship between gene polymorphism and platelet reactivity in patients treated with clopidogrelObjective: To investigate the relationship between platelet reactivity and gene polymorphism of CYP2C19*2,*3,ABCB1(C3435T),CYP3A4(IVS10+12G>A),CYP3A5(rs3800959)in patients after clopidogrel administration.Methods: Patients of Han population which met the inclusion criteria were included from June 2016 to March 2017,and were measured the maximum platelet aggregation rate.Meanwhile we analyzed their gene types of CYP2C19*2,*3,ABCB1(C3435T),CYP3A4(IVS10+12G>A),CYP3A5(rs3800959)and explored their relationship with high platelet reactivity(HPR)and low platelet reactivity(LPR).Results: 48 patients were included.The overall incidence of HPR was 25%(12/48).Mean maximum platelet aggregation rate was(56.21+2.94)%.The incidence of LPR was 75%(36/48).Mean maximum platelet aggregation rate was(34.48+11.64)%.There was no significant difference between HPR group and LPR group in general clinical data and drug use during hospitalization.CYP2C19*2: In group HPR,the GG,GA,AA genotypes were: 1 cases(8.33%),4 cases(33.33%),7 cases(58.33%).The frequency of allele G was 25%,A was 75%.In group LPR,the GG,GA,AA genotypes were: 13 cases(36.11%),17 cases(47.22%),6 cases(16.67%).The frequency of allele G was 59.72%,A was 40.28%.There was a significant difference in the frequency of AA group and allele A in HPR group and in LPR group(P=0.015,P=0.003).CYP2C19*3: Both in HPR group and LPR group,only GG type was detected.ABCB1(C3435T): In group HPR,the CC,CT,TT genotypes were: 0 cases(0%),7 cases(58.33%),5 cases(41.67%).The frequency of allele C was 29.17%,T was 70.83%.In group LPR,the CC,CT,TT genotypes were: 22 cases(61.11%),14 cases(38.89%),0 cases(0%).The frequency of allele C was 80.56%,T was 19.44%.There was a significant difference in the frequency of CC group,TT group and allele T in HPR group and in LPR group(P<0.001,P<0.001,P<0.001).CYP3A4(IVS10+12G>A): In group HPR,the GG,GA,AA genotypes were: 9 cases(75%),3 cases(25%),0cases(0%).The frequency of allele G was 87.5%,A was 12.5%.In group LPR,the GG,GA,AA genotypes were: 14 cases(38.89%),20 cases(55.56%),2cases(5.56%).The frequency of allele G was 66.67%,A was 33.33%.There was a significant difference in the frequency of GG group and allele A in HPR group and in LPR group(P=0.030,P=0.049).CYP3A5(rs3800959): In group HPR,the TT,TC,CC genotypes were: 9 cases(75%),2 cases(16.67%),1 cases(8.33%).The frequency of allele T was 83.33%,C was 16.67%.In group LPR,the TT,TC,CC genotypes were: 26 cases(72.22%),10 cases(27.78%),0 cases(0%).The frequency of allele T was 86.11%,C was 13.89%.There was no significant difference in the frequency of each genotype in HPR group and LPR group.Conclusion: Different platelet reactivities maybe were related to the mutations of genes of CYP2C19*2,ABCB1(C3435T),CYP3A4(IVS10+12G>A)after clopidogrel,but due to the limitations of the sample size,the results need further verification.Part 3: Investigation on the relationship between the use of clopidogrel and gene polymorphism in patients and medical workersObjective: To investigate the cognitive status of clopidogrel between medical workers and patients,and promote the rational use of drugs.Methods: We sent questionnaires randomly to medical workers at all levels of the hospital and in the Department of Cardiology of the Affiliated Hospital of Southwest Medical University.Through these two forms,we could analyse the cognitive status of clopidogrel of medical workers and patients.Then according to the results,we used the SPSS software for statistical analysis.Results: Among the 354 medical workers,there were 155 male and 199 female.The largest proportion(39.27%)come from East China.Different regions,ages,jobs,titles,educations,monthly incomes can affect the medical staff's cognition of basic drug information,clinical usage,contraindications,adverse reactions and metabolism related genes on the clopidogrel.In patients,the cognitive level of clopidogrel was generally low.The results showed that only 8 of the patients in the Department of Cardiology had heard of the drug,and the awareness rate was only about 4%.Conclusion: The medical staff should strengthen the knowledge updating and popularization of clopidogrel,and increase in patients of drug propaganda,training and education.To renew the medical personnel(including doctors,nurses and pharmacy technicians)of the clinical medication knowledge,and promote the application of better and more rational drug dispensing service. |
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