Thrombosis Research 135 (2015) 415–416
Contents lists available at ScienceDirect
Thrombosis Research journal homepage: www.elsevier.com/locate/thromres
Letter to the Editor-in-Chief Lack of association between Factor V Hong Kong and Venous thrombosis in the Chinese population
Dear Editors, Venous thrombosis (VT),which is a major medical disorder throughout the world,has a high morbidity and mortality [1]. VT is a multifactorial disease with both established environmental and genetic risk factors. Numerous genetic risk factors of VT have being found,including some genetic risk factors in the Chinese population [2]. However, the underlying molecular mechanisms of many inherited thrombotic events remain unsolved. Among the known genetic factors, the mutations located in F5 (for example FV Leiden) have a significant influence on Caucasians [3]. Several years ago,a mutation located in F5 was identified in Hong Kong Chinese, which resulted in p.Arg306Gly substitution. At that time, this mutation was found in 2 of 43 unrelated Hong Kong Chinese patients with a definite DVT history and 1 of 40 nonthrombotic patients. The authors showed the harm of this mutation by some experiments [4]. However,multiple studies in other countries failed to clear the risk of this mutation [5]. In order to determine whether FV Hong Kong is a genetic risk factor for VT in the Chinese population,we analysed this mutation in a large sample size. A total of 2638 unrelated Chinese participants were enrolled and analyzed, including 1334 healthy controls and 1304 patients with venous thrombosis. All of the cases and controls were described previously (Tang et al., 2013).Thrombosis was confirmed by objective investigations such as color Doppler ultrasonography and/or CT angiography. Informed consent was obtained from all participants, and the study was approved by the Ethics Committee of Union Hospital affiliated with Huazhong University of Science and Technology. This mutation was detected in all of the cases and controls by employing a PCR-restriction fragment length polymorphism (PCR-RFLP) method. The primer sequences for amplification were 5’- CTGGATGCCACTTAAAGA-3’ and 5’-GTTTGTAGAGTTGAAGCG-3’. The PCR product was a 623-bp fragment. The wild type PCR product can be digested by the restrictive enzyme Mva I,but the mutant type can't. Digestion was carried out by incubating 10 ul of the PCR product in 17 ul double distilled water with 3 ul of 10 x buffer B and 5 U FastDigest® MvaI (Fermentas, Burlington, Canada) for 10 min at 37 °C. The digestion products were separated by 1.5% agarose gel electrophoresis. Altogether, 96 DNA samples were selected and subjected to sequencing to verify the genotyping results determined with the PCR-RFLP assays. The genotyping data from the PCR-RFLP assays were completely consistent with those obtained by sequencing. We found 7 heterozygotes in the VT group and 15 heterozygotes in the control group. None of these participants is homozygous mutant (Table 1).
http://dx.doi.org/10.1016/j.thromres.2014.11.013 0049-3848/© 2014 Elsevier Ltd. All rights reserved.
The P value is 0.097 and the Odds ratio is 0.475.This result shows that FV Hong Kong is not a risk factor for venous thrombosis in the Chinese population. Factor V has two kinds of role in coagulation process. On one hand, activated factor V (FVa) functions as a cofactor to factor Xa (FXa) in the activation of prothrombin, enhancing the rate of thrombin generation several thousand times. The procoagulant function of FVa is down-regulated by the anticoagulant serine protease activated protein C (APC). APC cleaves FVa at 3 sites-Arg306, Arg506, and Arg679-which possible results in the loss of FVa activity. On the other hand, circulating FV has an anticoagulant role functioning as a synergistic APC cofactor to protein S in the inactivation of FVIIIa [6]. The mutation of FV Hong Kong delay the APCmediated inactivation of the FVa, partly promote the coagulation progress;however,this mutation is able to support APC-mediated degradation of FVIIIa [7]. FV Hong Kong is not associated with increased risk of thrombosis [8]. In conclusion, our results indicate that FV Hong Kong is not a genetic risk factor for venous thrombosis in the Chinese population.
Funding This study was supported by grants from the National Natural Sciences Foundation of China (No. 81370622 and No. 81400099) and the Independent Innovation Foundation of Huazhong University of Science and Technology (No. 01-18-530045, 2013QN213).
Conflict of Interest We declare that we have no conflict of interest.
Table 1 Factor V Hong Kong in the 2638 Chinese population. Factor V Hong Kong
Control group n = 1334
Venous thrombosis n = 1304
Age(yr,mean) Male,n(%) AA AG GG
51.3 ± 14.2 48.7% 1319(98.9%) 15(1.1%) 0(0%)
51.7 ± 13.8 48.9% 1297(99.5%) 7(0.5%) 0(0%)
dbSNP ID: rs118203905; P value 0.097; chi2 = 2.75; Odds ratio = 0.475. dbSNP, single nucleotide polymorphism database of the National Center for Biotechnology Information. (http://www.ncbi.nlm.nih.gov/projects/SNP). Comparisons between the controls and each case group were assessed with the use of the chi-square test. A two tailed P b 0.05 was considered significant.
416
Letter to the Editor-in-Chief
References [1] Flinterman LE, van Hylckama Vlieg A, Cannegieter SC, Rosendaal FR. Long-term survival in a large cohort of patients with venous thrombosis: Incidence and predictors. PLoS Med Jan 2012;9(1):e1001155. [2] Tang L, Wang HF, Lu X, Jian XR, Jin B, Zheng H, et al. Common genetic risk factors for venous thrombosis in the Chinese population. Am J Hum Genet Feb 7 2013;92(2): 177–87. [3] Seligsohn U, Lubetsky A. Genetic susceptibility to venous thrombosis. N Engl J Med Apr 19 2001;344(16):1222–31. [4] Chan WP, Lee CK, Kwong YL, Lam CK, Liang R. A Novel Mutation of Arg306 of Factor V Gene in Hong Kong Chinese. Blood Feb 15 1998;91(4):1135–9. [5] Dölek B, Eraslan S, Eroğlu S, Kesim BE, Ulutin T, Yalçiner A, et al. Molecular Analysis of Factor V Leiden, Factor V Hong Kong, Factor II G20210A, Methylenetetrahydrofolate Reductase C677T, and A1298C Mutations Related to Turkish Thrombosis Patients. Clin Appl Thromb Hemost Oct 2007;13(4):435–8. [6] Dahlbäck B, Hildebrand B. Inherited resistance to activated protein C is corrected by anticoagulant cofactor activity found to be a property of factor V. Proc Natl Acad Sci U S A Feb 15 1994;91(4):1396–400. [7] Norstrøm E, Thorelli E, Dahlbäck B. Functional characterization of recombinant FV Hong Kong and FV Cambridge. Blood Jul 15 2002;100(2):524–30. [8] Liang R, Lee CK, Wat MS, Kwong YL, Lam CK, Liu HW. Clinical significance of Arg306 mutations of factor V gene. Blood 1998;92:2599–600.
Zhi-Peng Cheng1 Liang Tang1 Institute of Hematology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China Collaborative Innovation Center of Hematology, Huazhong University of Science and Technology, Wuhan, China
Hui Liu1 Institute of Hematology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
1
Equal contribution.
Wei Zeng Qing-Yun Wang Institute of Hematology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China Collaborative Innovation Center of Hematology, Huazhong University of Science and Technology, Wuhan, China Ying-Ying Wu Institute of Hematology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China Bei Hu Yu Hu⁎ Institute of Hematology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China Collaborative Innovation Center of Hematology, Huazhong University of Science and Technology, Wuhan, China ⁎Corresponding author at: Institute of Hematology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei 430022, China. Tel.: +86 27 85726335; fax: +86 27 85726387. E-mail address: [email protected]. 21 October 2014
Contents lists available at ScienceDirect
Thrombosis Research journal homepage: www.elsevier.com/locate/thromres
Letter to the Editor-in-Chief Lack of association between Factor V Hong Kong and Venous thrombosis in the Chinese population
Dear Editors, Venous thrombosis (VT),which is a major medical disorder throughout the world,has a high morbidity and mortality [1]. VT is a multifactorial disease with both established environmental and genetic risk factors. Numerous genetic risk factors of VT have being found,including some genetic risk factors in the Chinese population [2]. However, the underlying molecular mechanisms of many inherited thrombotic events remain unsolved. Among the known genetic factors, the mutations located in F5 (for example FV Leiden) have a significant influence on Caucasians [3]. Several years ago,a mutation located in F5 was identified in Hong Kong Chinese, which resulted in p.Arg306Gly substitution. At that time, this mutation was found in 2 of 43 unrelated Hong Kong Chinese patients with a definite DVT history and 1 of 40 nonthrombotic patients. The authors showed the harm of this mutation by some experiments [4]. However,multiple studies in other countries failed to clear the risk of this mutation [5]. In order to determine whether FV Hong Kong is a genetic risk factor for VT in the Chinese population,we analysed this mutation in a large sample size. A total of 2638 unrelated Chinese participants were enrolled and analyzed, including 1334 healthy controls and 1304 patients with venous thrombosis. All of the cases and controls were described previously (Tang et al., 2013).Thrombosis was confirmed by objective investigations such as color Doppler ultrasonography and/or CT angiography. Informed consent was obtained from all participants, and the study was approved by the Ethics Committee of Union Hospital affiliated with Huazhong University of Science and Technology. This mutation was detected in all of the cases and controls by employing a PCR-restriction fragment length polymorphism (PCR-RFLP) method. The primer sequences for amplification were 5’- CTGGATGCCACTTAAAGA-3’ and 5’-GTTTGTAGAGTTGAAGCG-3’. The PCR product was a 623-bp fragment. The wild type PCR product can be digested by the restrictive enzyme Mva I,but the mutant type can't. Digestion was carried out by incubating 10 ul of the PCR product in 17 ul double distilled water with 3 ul of 10 x buffer B and 5 U FastDigest® MvaI (Fermentas, Burlington, Canada) for 10 min at 37 °C. The digestion products were separated by 1.5% agarose gel electrophoresis. Altogether, 96 DNA samples were selected and subjected to sequencing to verify the genotyping results determined with the PCR-RFLP assays. The genotyping data from the PCR-RFLP assays were completely consistent with those obtained by sequencing. We found 7 heterozygotes in the VT group and 15 heterozygotes in the control group. None of these participants is homozygous mutant (Table 1).
http://dx.doi.org/10.1016/j.thromres.2014.11.013 0049-3848/© 2014 Elsevier Ltd. All rights reserved.
The P value is 0.097 and the Odds ratio is 0.475.This result shows that FV Hong Kong is not a risk factor for venous thrombosis in the Chinese population. Factor V has two kinds of role in coagulation process. On one hand, activated factor V (FVa) functions as a cofactor to factor Xa (FXa) in the activation of prothrombin, enhancing the rate of thrombin generation several thousand times. The procoagulant function of FVa is down-regulated by the anticoagulant serine protease activated protein C (APC). APC cleaves FVa at 3 sites-Arg306, Arg506, and Arg679-which possible results in the loss of FVa activity. On the other hand, circulating FV has an anticoagulant role functioning as a synergistic APC cofactor to protein S in the inactivation of FVIIIa [6]. The mutation of FV Hong Kong delay the APCmediated inactivation of the FVa, partly promote the coagulation progress;however,this mutation is able to support APC-mediated degradation of FVIIIa [7]. FV Hong Kong is not associated with increased risk of thrombosis [8]. In conclusion, our results indicate that FV Hong Kong is not a genetic risk factor for venous thrombosis in the Chinese population.
Funding This study was supported by grants from the National Natural Sciences Foundation of China (No. 81370622 and No. 81400099) and the Independent Innovation Foundation of Huazhong University of Science and Technology (No. 01-18-530045, 2013QN213).
Conflict of Interest We declare that we have no conflict of interest.
Table 1 Factor V Hong Kong in the 2638 Chinese population. Factor V Hong Kong
Control group n = 1334
Venous thrombosis n = 1304
Age(yr,mean) Male,n(%) AA AG GG
51.3 ± 14.2 48.7% 1319(98.9%) 15(1.1%) 0(0%)
51.7 ± 13.8 48.9% 1297(99.5%) 7(0.5%) 0(0%)
dbSNP ID: rs118203905; P value 0.097; chi2 = 2.75; Odds ratio = 0.475. dbSNP, single nucleotide polymorphism database of the National Center for Biotechnology Information. (http://www.ncbi.nlm.nih.gov/projects/SNP). Comparisons between the controls and each case group were assessed with the use of the chi-square test. A two tailed P b 0.05 was considered significant.
416
Letter to the Editor-in-Chief
References [1] Flinterman LE, van Hylckama Vlieg A, Cannegieter SC, Rosendaal FR. Long-term survival in a large cohort of patients with venous thrombosis: Incidence and predictors. PLoS Med Jan 2012;9(1):e1001155. [2] Tang L, Wang HF, Lu X, Jian XR, Jin B, Zheng H, et al. Common genetic risk factors for venous thrombosis in the Chinese population. Am J Hum Genet Feb 7 2013;92(2): 177–87. [3] Seligsohn U, Lubetsky A. Genetic susceptibility to venous thrombosis. N Engl J Med Apr 19 2001;344(16):1222–31. [4] Chan WP, Lee CK, Kwong YL, Lam CK, Liang R. A Novel Mutation of Arg306 of Factor V Gene in Hong Kong Chinese. Blood Feb 15 1998;91(4):1135–9. [5] Dölek B, Eraslan S, Eroğlu S, Kesim BE, Ulutin T, Yalçiner A, et al. Molecular Analysis of Factor V Leiden, Factor V Hong Kong, Factor II G20210A, Methylenetetrahydrofolate Reductase C677T, and A1298C Mutations Related to Turkish Thrombosis Patients. Clin Appl Thromb Hemost Oct 2007;13(4):435–8. [6] Dahlbäck B, Hildebrand B. Inherited resistance to activated protein C is corrected by anticoagulant cofactor activity found to be a property of factor V. Proc Natl Acad Sci U S A Feb 15 1994;91(4):1396–400. [7] Norstrøm E, Thorelli E, Dahlbäck B. Functional characterization of recombinant FV Hong Kong and FV Cambridge. Blood Jul 15 2002;100(2):524–30. [8] Liang R, Lee CK, Wat MS, Kwong YL, Lam CK, Liu HW. Clinical significance of Arg306 mutations of factor V gene. Blood 1998;92:2599–600.
Zhi-Peng Cheng1 Liang Tang1 Institute of Hematology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China Collaborative Innovation Center of Hematology, Huazhong University of Science and Technology, Wuhan, China
Hui Liu1 Institute of Hematology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
1
Equal contribution.
Wei Zeng Qing-Yun Wang Institute of Hematology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China Collaborative Innovation Center of Hematology, Huazhong University of Science and Technology, Wuhan, China Ying-Ying Wu Institute of Hematology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China Bei Hu Yu Hu⁎ Institute of Hematology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China Collaborative Innovation Center of Hematology, Huazhong University of Science and Technology, Wuhan, China ⁎Corresponding author at: Institute of Hematology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei 430022, China. Tel.: +86 27 85726335; fax: +86 27 85726387. E-mail address: [email protected]. 21 October 2014
