|Year : 2013 | Volume
| Issue : 3 | Page : 97-101
Detection of Janus kinase 2 V617F mutation in healthy cigarette smokers
Samy B.M. El-Hady1, Amina M. Elnaggar1, Eman Almasry2
1 Department of Clinical Pathology, Faculty of Medicine, Zagazig University, Zagazig, Egypt
2 Department of Biochemistry, Faculty of Medicine, AlMenia University, Almenia, Egypt
|Date of Submission||18-Dec-2012|
|Date of Acceptance||27-Feb-2012|
|Date of Web Publication||19-Jun-2014|
Samy B.M. El-Hady
MD, Department of Clinical Pathology, Faculty of Medicine, Zagazig University, Zagazig
Source of Support: None, Conflict of Interest: None
Janus kinases are cytoplasmic tyrosine kinases that mediate signaling from the cytokine receptors to the cell nucleus. Janus kinase 2 mutation (JAK2 V617F) analysis has been endorsed by the WHO for diagnosing polycythemia vera, essential thrombocythemia, and primary myelofibrosis. The aim of this study was to assess JAK2 V617F point mutation in healthy cigarette smokers compared with healthy nonsmokers and to correlate the presence of this mutation with some clinical and laboratory variables.
Materials and methods
Group I comprised 34 cigarette smokers who have been smoking 10 or more cigarettes per day, every day of the week, for at least 10 consecutive years. Group II comprised 42 men who were nonsmokers with no history of drug abuse. In addition to routine laboratory investigations, detection of JAK2 V617F point mutation in peripheral blood neutrophils was assessed for all participants.
In this study, we found an increased percentage of JAK2 V617F mutation in cigarette smokers compared with nonsmokers. Further, we found a significant positive correlation between the percentage of JAK2 V617F mutation and age in both groups.
JAK2 V617F mutation has been detected in the healthy population; however, its incidence significantly increases in cigarette smokers. The mechanisms leading to excess JAK2 mutation and the importance of this mutation in smokers are yet to be elucidated and an adequate follow-up of healthy individuals who carry the mutation is recommended.
Keywords: cigarette smokers, JAK2 V617F, Janus kinase 2 mutation
|How to cite this article:|
El-Hady SB, Elnaggar AM, Almasry E. Detection of Janus kinase 2 V617F mutation in healthy cigarette smokers. Egypt J Haematol 2013;38:97-101
|How to cite this URL:|
El-Hady SB, Elnaggar AM, Almasry E. Detection of Janus kinase 2 V617F mutation in healthy cigarette smokers. Egypt J Haematol [serial online] 2013 [cited 2019 Aug 23];38:97-101. Available from: http://www.ehj.eg.net/text.asp?2013/38/3/97/128297
| Introduction|| |
Polycythemia can result from polycythemia vera (PV) or be secondary to tissue hypo-oxygenation, resulting from high altitude 1, alveolar hypoventilation 2, or cigarette smoking 3–5. PV is a chronic myeloproliferative disorder characterized by an increased red cell mass. Thrombosis and hemorrhage are the major causes of morbidity and mortality in PV. Some patients may develop myelofibrosis (MF) or acute myeloid leukemia. The Janus kinase 2 (JAK2)/signal transducers and activators of transcription (STAT) pathway influences cell proliferation, activation, and apoptosis 6–8. JAK2 is a tyrosine kinase that is associated with the cytoplasmic domain of the erythropoietin (Epo) receptor. It transduces signals, especially those triggered by hematopoietic growth factors, such as Epo 9. When Epo binds to the Epo receptor, dimerization of receptor subunits occurs. Hence, two JAKs are brought in proximity and undergo transphosphorylation. The activated JAKs subsequently phosphorylate their major substrates, STATs, which once activated enter the cell nucleus and bind to specific regulatory sequences to activate or repress the transcription of target genes. Thus, the JAK/STAT cascade provides a direct mechanism for translating an extracellular signal into a transcriptional response 7. A point mutation in the Janus kinase 2 gene was identified in several myeloproliferative neoplasms, 10–13 most frequently in PV (65–97%), essential thrombocytosis (ET) (23–57%), and MF (35–57%). It is also infrequently present in myelodysplastic syndrome, chronic myelomonocytic leukemia, and other atypical chronic myeloid disorders 14. The common occurrence of this mutation in BCR/ABL-negative myeloproliferative neoplasms can be used as a unique molecular marker for distinguishing PV, ET, and MF from reactive hematopoietic disorders 15. Coexistence of JAK2 V617F mutation and BCR-ABL rearrangement is rare but has been reported 16. The V617F mutation is a G>T transversion at nucleotide 2343, resulting in the substitution of phenylalanine for valine (V617F) in the JAK2 protein. This point mutation appears to cause constitutive activation of the JAK2 tyrosine kinase owing to loss of autoinhibitory control 17. The mutant has enhanced kinase activity, and when overexpressed together with the Epo receptor in cells it causes hyperactivation of Epo-induced cell signaling. This may explain the hypersensitivity of PV progenitor cells to growth factors and cytokines. Observations suggest that this mutation occurs at the level of the common myeloid progenitor or upstream at the level of a lymphomyeloid multipotent progenitor cell 10,18. Identification of a JAK2 V617F mutation establishes the presence of a clonal disorder 19 and is an important diagnostic marker for these disorders 15. The mutation was not found in several reactive hematological states 20 and in people with high affinity hemoglobin 21. Some smokers develop erythrocytosis, whereas others do not 22,23. After smoking cessation, the hematocrit level usually decreases 23,24. Surprisingly, the exact mechanism linking smoking and erythrocytosis has not been studied extensively and was hypothesized to be related to decreased tissue oxygenation. This in turn may cause increased Epo secretion from the kidneys 25. The cause for decreased tissue oxygenation was hypothesized to be increased levels of carboxy-hemoglobin 26. When carboxy-hemoglobin levels increase, the hemoglobin–oxygen dissociation curve is shifted to the left, resulting in greater affinity of hemoglobin to oxygen, which in turn is not released into the tissue 27. Accordingly, the state of erythrocytosis may in fact be an adaptation mechanism to the hypoxia induced by carbon monoxide 5, 26, 28.
This study had two main objectives. The first was to examine the effect of smoking on the JAK2 mutation percentage in the healthy population. The second objective was to examine a possible correlation between this mutation and some clinical and laboratory variables.
| Subjects and methods|| |
This study included 76 men chosen from blood bank donors. They were free from any disease and had no history of drug abuse. They were not frequent blood donors. They were categorized into two groups.
Group I: Group I comprised 34 men with a mean age of 37.1±7.8 years. They were only cigarette smokers and had been smoking 10 or more cigarettes per day, every day of the week, for at least 10 consecutive years. The duration of smoking ranged from 12 to 23 years.
Group II: Group II comprised 42 men with a mean age of 36.2±9.6 years who were nonsmokers.
All individuals were subjected to the following:
- Full history taking and complete clinical examination.
- Laboratory investigations including:
- Complete blood count using Sysmex SF 3000 (Kobe, Japan), with examination of Leishman-stained films.
- Liver and kidney function tests using a Roche Hitachi 902 Automatic Analyzer (Roche Diagnostics GmbH, Mannheim, Germany).
- Analysis of serum ferritin level using Cobas e411-Roche Diagnostics.
- Special investigations, which included:
- Detection of Epo in plasma by an enzyme-linked immunosorbent assay-based technique (supplied by R&D Systems Inc., Minneapolis, Minnesota, USA).
- JAK2 V617F mutation assay, comprising:
- Neutrophil isolation: In this step, peripheral blood was collected using EDTA as an anticoagulant. Samples were processed on the same day. Neutrophils were isolated by dextran (Fisher Scientific, Pittsburgh, Pennsylvania, USA) sedimentation of RBC and subsequent centrifugation of plasma on a Ficoll-Hypaque density gradient (GE Healthcare Life Sciences, Uppsala, Sweden) at 400g. Contaminating erythrocytes in the sedimented cell pellet were removed by hypotonic lysis. Thereafter, neutrophils were washed three times with PBS (Sigma Immunochemicals, St Louis, Missouri, USA). Final cell suspension was more than 97% pure.
- DNA extraction (Wizard Genomic DNA Purification Kit; Promega, Madison, Wisconsin, USA): As the first step in the purification procedure, white blood cells and their nuclei are lysed using a nuclei lysis solution. The cellular proteins are then removed by a salt precipitation step, which precipitates the proteins but leaves the high-molecular-weight genomic DNA in solution. Finally, the genomic DNA is concentrated and desalted by isopropanol precipitation. DNA is rehydrated and stored at 2–8°C.
- JAK2 MutaSearch Assay (allele-specific PCR): The JAK2 MutaSearch assay (Ipsogen; Luminy Biotech, Marseille, France), which detects JAK2 wild type and V617F alleles in genomic DNA, was used according to the manufacturer’s protocol. Reactions were performed using the following PCR program: 95°C for 10 min, followed by 50 cycles of 95°C for 15 s and 66°C for 1 min. A Light Cycler instrument was used (Roche diagnostics). The Cp value for each sample (controls, cutoff sample ‘cos’, and unknown samples) was calculated. For all samples, the &Dgr;Cp value was ascertained by calculating the difference between the CpVF (mean Cp value obtained with PPM-Jak2 V617F) and CpWT (mean Cp value obtained with PPM-Jak2 WT). The grey zone of the test has been defined as +/−7% of the &Dgr;CpCOS For each unknown sample and control the &Dgr;&Dgr;Cp value was calculated by subtracting &Dgr;Cpsample from &Dgr;Cpcos If the &Dgr;&Dgr;Cp value of the sample is below the &Dgr;Cp value of the cutoff sample minus 7%, the sample is considered negative (JAK2 V617F mutation not detected) and if the &Dgr;&Dgr;Cp value of the sample is above the &Dgr;Cp value of the cutoff sample plus 7% the sample is considered positive (JAK2 V617F mutation is detected).
Laboratory results were expressed as mean±SD (X±SD) and compared using the two-sample t-test. The percentage ofpatients positive for JAK2 V617F was determined. Correlation analysis was performed with Pearson’s correlation test. P values below 0.05 were considered significant. Data were tabulated statistically and analyzed using SPSS version 20.0 for windows (SPSS Inc., Chicago, Illinois, USA).
| Results|| |
None of the participants in this cohort had known hematological, liver, or kidney diseases. Some laboratory data of the healthy male cigarette smokers (group I) and nonsmokers (group II) are presented in [Table 1].
[Table 2] and [Figure 1] show that the percentage of individuals with JAK 2 mutation was 20.6% in group I and 9.5% in group II. [Table 3] shows that there were significant positive correlations between JAK 2 mutation and age in both groups (r=0.48, P<0.05 in group I, r=0.47, P<0.05 in group II).
|Table 2: Comparison between the studied groups as regards JAK2 V617F mutation|
Click here to view
|Table 3: Correlations (r) between JAK2 V617F mutation and laboratory findings in the studied groups|
Click here to view
| Discussion|| |
A somatic mutation of the Janus kinase 2 gene is present in most patients with PV and in about half of those with essential thrombocythemia and chronic idiopathic MF 29. The JAK2 V617F mutation confers cytokine-independent growth and cytokine hypersensitivity of cell lines 10. Targeted drugs are becoming available for treatment and their use will necessitate evaluation of the JAK2 V617F allele burden at diagnosis and for subsequent patient monitoring. Ruxolitinib (Jakafi; Jakavi) is one of the Janus kinase inhibitors that was approved by the USFDA for treatment of intermediate-risk or high-risk MF 30,31. As per 2008 WHO criteria and diagnostic algorithms, 32 peripheral blood JAK2 V617F screening is currently the preferred initial test for evaluating patients with suspected PV. Not all clonal cells express the JAK2 V617F point mutation, and granulocyte enrichment has been suggested for mutation detection assays 33. The aim of this study was to evaluate the presence of JAK2 V617F mutation in healthy individuals, smokers, and nonsmokers and assess its relation to different clinical variables. In our analysis, JAK2 V617F mutation was found in healthy individuals but its percentage is higher in smokers than in nonsmokers. This finding is in agreement with the results of the study by Weinberg et al. 34, who found greater prevalence of the mutation in smokers than in nonsmokers. Many studies describe JAK2 V617F mutation in healthy individuals and in people with nonhematologic conditions 35,36. Also, Sidon et al. 37 detected a low level of the mutation in 10% of the control samples in their study. In two other studies, the JAK2 V617F mutant was found in normal control samples 38,39. Similarly, in their study on allogeneic stem cell transplant recipients with MF, Krger et al. 40 detected the mutation in 7% of healthy control individuals. In contrast, all the 10 normal control samples in the study by Cankovic et al. 33 were negative for the mutation. Similarly, James et al. 19 did not find the mutation in any of the five individuals with secondary erythrocytosis. Yet, it is possible that both groups were too small to be representative of a much larger population. Also, Mast et al. 4 did not find the mutation among 138 high-frequency blood donors, and Tefferi et al. 41 did not find the mutation in granulocytes of 19 secondary polycytemia patients, nor in the 11 healthy individuals without secondary polycythemia. Similarly, Passamonti et al. 20 did not find the mutation in any of 75 healthy individuals or in the 89 individuals with reactive hematological conditions. Finally, McClure et al. 21 failed to find the mutation in any of 22 patients with high oxygen-affinity hemoglobin using the PCR method. Weinberg et al. 34 suggested that the difference in the prevalence of JAK2 mutation reported in various populations may be partly dependent on the sensitivity of the method used to detect the mutation, the criteria used to select the study populations, and the number of participants. Also, it may be attributed to the cells used for DNA extraction because not all clonal cells express the JAK2 V617F point mutation 33.
We suggest that the significant increase in JAK2 mutation prevalence in the smokers in our study may be due to poor DNA repair. Erythrocytosis in smokers may lead to more DNA repair errors secondary to enhanced cell division 35, 42, 43 and thus to an increased prevalence of the JAK2 mutation. However, in our study, we cannot find a significant correlation between the mutation and erythrocytosis, and further studies are needed to establish this hypothesis. Sidon et al. 37 hypothesized that discovering the mutation in healthy individuals raised the possibility that the mutation is harbored before the occurrence of the chronic myeloproliferative disorder. This should be confirmed by an adequate follow-up of healthy individuals who carry the mutation. However, the annual incidence of PV and essential thrombocythemia is very low, ranging from 2 to 2.5 per 100 000 individuals, and the JAK2 V617F mutation occurs in healthy individuals at a higher frequency than expected in the early detection of clinical chronic myeloproliferative disorders. One hypothesis put forth by Passamonti et al. 20 to explain JAK2 V617F in healthy individuals is that the first progenitor to harbor JAK2 V617F might be more differentiated in healthy individuals than in patients with a chronic myeloproliferative disorder. At this stage, the self-renewal potential and the capacity of the mutated cell to differentiate are decreased and the clone it generates might be prone to death. In this hypothesis, the mutated clone does not generate a clinical phenotype and it may disappear at serial evaluation. In contrast, in PV and in MF, the JAK2 V617F mutation is harbored in a lymphomyeloid progenitor cell 44. Also, studies suggest that gene dosage might have a role in determining whether a patient will develop PV, MF, or ET. Approximately 25% of patients with PV have been shown to be homozygous for the JAK2 V617F allele, whereas most patients with ET are heterozygous or have the wild type. Homozygosity increases the expression of the mutated protein and eliminates the competition with wild-type protein 45,46. A report demonstrated a correlation between hematologic improvement and a reduction in the proportion of the JAK2 V617F mutant alleles 47.
In our study, we could not find a significant correlation between JAK2 mutation prevalence and the degree of erythrocytosis, hematocrit, white blood cell count, platelet count, or Epo level. Our findings agree with those of Weinberg et al. 34, who did not find a correlation between JAK2 mutation prevalence or frequency and the degree of erythrocytosis, hematocrit, or white blood cell count. Hematocrit was higher in smokers, yet Epo levels were normal. One would expect a higher level of Epo in smokers as an explanation for their elevated hematocrit. This may in fact point to relatively increased erythrocytosis in smokers with resultant downregulation of Epo secretion. Of particular interest is the relation between JAK2 mutation and age. DNA repair mechanisms are known to become impaired with age 48. Xu et al. 38 found a higher prevalence of JAK2 mutation in individuals who were 13.6 years older than the sample average. In contrast, in an analysis of 198 individuals, Martinaud et al. 36 did not find a difference in mutation rate between individuals older than 40 years and those younger than 40 years.
| Conclusion|| |
The mechanism leading to excess JAK2 mutation and the importance of this mutation in smokers are yet to be elucidated and an adequate follow-up of healthy individuals who carry the mutation is recommended.
| References|| |
|1.||Frisancho A.Functional adaptation to high altitude hypoxia.Science1975;187:313–319. |
|2.||Auchincloss J, Cook E, Renzetti A.Clinical and physiological aspects of a case of obesity, polycythemia and alveolar hypoventilation.J Clin Invest1955;34:1537–1545. |
|3.||Ernst E, Matrai A, Schmuz C, Magyarosy I.Dose-effect relationship between smoking and blood rheology.Br J Haematol1987;65:485–487. |
|4.||Mast A, Foster T, Pinder H, Beczkiewicz C, Bellissimo D, Murphy A, et al..Behavioral, biochemical, and genetic analysis of iron metabolism in high-intensity blood donors.Transfusion2008;48:2197–2204. |
|5.||Spiers A, Levine M.Smokers’ polycythaemia.Lancet1983;1:120. |
|6.||Tefferi A, Pardanani A.Evaluation of ‘increased’ hemoglobin in the JAK2 mutations era: a diagnostic algorithm based on genetic tests.Mayo Clin Proc2007;82:599–604. |
|7.||Rawlings J, Rosler K, Harrison D.The JAK/STAT signaling pathway.J Cell Sci2004;117:1281–1283. |
|8.||Ihle J, Gilliland D.Jak2: normal function and role in hematopoietic disorders.Curr Opin Genet Dev2007;17:8–14. |
|9.||Tefferi A.JAK2 mutations in polycythemia vera-molecular mechanisms and clinical applications.N Engl J Med2007;356:444–445. |
|10.||Baxter E, Scott L, Campbell P, East C, Fourouclas N, Swanton S, et al..Acquired mutation of the tyrosine kinase JAK2 in human myeloproliferative diseases.Lancet2005;365:1054–1061. |
|11.||James C, Ugo V, LeCouedic J, Staerk J, Delhommeau F, Lacout C, et al..A unique clonal JAK2 mutation leading to constitutive signaling causes polycythaemia vera.Nature2005;434:1144–1148. |
|12.||Kralovics R, Passamonti F, Buser A, Teo S, Tiedt R, Passweg J, et al..A gain-of function mutation of JAK2 in myeloproliferative disorders.N Engl J Med2005;352:1779–1790. |
|13.||Levine R, Wadleigh M, Cools J, Ebert B, Wernig G, Huntly B, et al..Activating mutation of the tyrosine kinase JAK2 in polycythemia vera, essential thrombocythemia, and myeloid metaplasia with myelofibrosis.Cancer Cell2005;7:387–397. |
|14.||Steensma D, Dewald G, Lasho T, Powell H, McClure R, Levine R, et al..The JAK2 V617F activating tyrosine kinase mutation is an infrequent event in both ‘atypical’ myeloproliferative disorders and myelodysplastic syndromes.Blood2005;106:1207–1209. |
|15.||Campbell P, Scott L, Buck G, Wheatley K, East C, Marsden J, et al..Definition of subtypes of essential thrombocythaemia and relation to polycythaemia vera based on JAK2 V617F mutation status: a prospective study.Lancet2005;366:1945–1953. |
|16.||Pieri L, Spolverini A, Scappini B, Occhini U, Birtolo S, Bosi A, et al..Concomitant occurrence of BCR-ABL and JAK2 V617F mutation.Blood2011;118:3445–3446. |
|17.||Lindauer K, Loerting T, Liedl K, Kroemer R.Prediction of the structure of human Janus kinase 2 (JAK2) comprising the two carboxy-terminal domains reveals a mechanism for autoregulation.Protein Eng2001;14:27–37. |
|18.||Jamieson C, Gotlib J, Durocher J.The JAK2 V617F mutation occurs in hematopoietic stem cells in polycythemia vera and predisposes toward erythroid differentiation.Proc Natl Acad Sci USA2006;103:6224–6229. |
|19.||James C, Delhommeau F, Marzac C, Teyssandier I, Couédic J, Giraudier S, et al..Detection of JAK2 V617F as a first diagnostic test for erythrocytosis.Leukemia2006;20:350–353. |
|20.||Passamonti F, Rumi E, Pietra D, Lazzarino M, Cazzola M.JAK2 (V617F) mutation in healthy individuals.Br J Haematol2007;136:678–679. |
|21.||McClure R, Hoyer J, Mai M.The JAK2 V617F mutation is absent in patients with erythrocytosis due to high oxygen affinity hemoglobin variants.Hemoglobin2006;30:487–489. |
|22.||Tanabe N, Ohnishi K, Fukui H, Ohno R.Effect of smoking on the serum concentration of erythropoietin and granulocyte-colony stimulating factor.Intern Med1997;36:680–684. |
|23.||Van Tiel E, Peeters P, Smit H, Nagelkerke N, Van Loon A, Grobbee D, Bueno-de-Mesquita H.Quitting smoking may restore hematological characteristics within five years.Ann Epidemiol2002;12:378–388. |
|24.||Haustein K, Krause J, Haustein H, Rasmussen T, Cort N.Effects of cigarette smoking or nicotine replacement on cardiovascular risk factors and parameters of haemorheology.J Intern Med2002;252:130–139. |
|25.||Golde D, Hocking W, Koeffler H, Adamson J.Polycythemia: mechanisms and management.Ann Intern Med1981;95:71–87. |
|26.||Sagone J, Balcerzak S.Smoking as a cause of erythrocytosis.Ann Intern Med1975;82:512–515. |
|27.||Smith J, Landaw S.Smokers’ polycythemia.N Engl J Med1978;298:6–10. |
|28.||McAloon E, Streiff R, Kitchens C.Erythrocytosis associated with carboxyhemoglobinemia in smokers.South Med J1980;73:137–139. |
|29.||Passamonti F, Rumi E, Pietra D, Della Porta M, Boveri E, Pascutto C, et al..Relation between JAK2 (V617F) mutation status, granulocyte activation, and constitutive mobilization of CD34+ cells into peripheral blood in myeloproliferative disorders.Blood2006;107:3676–3682. |
|30.||Mesa R, Uma Y, Peter K.‘Ruxolitinib’.Nat Rev Drug Discov2012;11:103–104. |
|31.||Harrison C, Kiladjian J, Al-Ali H, Gisslinger H, Waltzman R, Stalbovskaya V, et al..‘JAK inhibition with ruxolitinib versus best available therapy for myelofibrosis’.N Engl J Med2012;366:787–798. |
|32.||Tefferi A, Vardiman J.Classification and diagnosis of myeloproliferative neoplasms: the 2008 World Health Organization criteria and point-of-care diagnostic algorithms.Leukemia2008;22:14–22. |
|33.||Cankovic M, Whiteley L, Hawley R, Zarbo R, Chitale D.Clinical performance of JAK2 V617F mutation detection assays in a molecular diagnostics laboratory. Evaluation of screening and quantitation methods.Am J Clin Pathol2009;132:713–721. |
|34.||Weinberg I, Borohovitz A, Krichevsky S, Perlman R, Ben-Yehuda A, Ben-Yehuda D.Janus kinase V617F mutation in cigarette smokers.Am J Hematol2012;87:5–8. |
|35.||Tagariello G, Di Gaetano R, Sartori R, Zanotto D, Belvini D, Radossi P, et al..The JAK2(V617F) tyrosine kinase mutation in blood donors with upper-limit haematocrit levels.Blood Transfus2009;7:111–116. |
|36.||Martinaud C, Brisou P, Mozziconacci M.Is the JAK2(V617F) mutation detectable in healthy volunteers?Am J Hematol2010;85:287–288. |
|37.||Sidon P, El Housni H, Dessars B, Heimann P.The JAK2V617F mutation is detectable at very low level in peripheral blood of healthy donors.Leukemia2006;20:1622. |
|38.||Xu X, Zhang Q, Luo J, Xing S, Li Q, Krantz S, et al..JAK2(V617F): prevalence in a large Chinese hospital population.Blood2007;109:339–342. |
|39.||Chen Q, Lu P, Jones AV, Cross N, Silver R, Wang Y.Amplification refractory mutation system, a highly sensitive and simple polymerase chain reaction assay, for the detection of JAK2 V617F mutation in chronic myeloproliferative disorders.J Mol Diagn2007;9:272–276. |
|40.||Krger N, Badbaran A, Holler E, Hahn J, Kobbe G, Bornhäuser M, et al..Monitoring of the JAK2-V617F mutation by highly sensitive quantitative real-time PCR after allogeneic stem cell transplantation in patients with myelofibrosis.Blood2007;109:1316–1321. |
|41.||Tefferi A, Sirhan S, Lasho T, Schwager S, Li C, Dingli D, et al..Concomitant neutrophil JAK2 mutation screening and PRV-1 expression analysis in myeloproliferative disorders and secondary polycythaemia.Br J Haematol2005;131:166–171. |
|42.||Preston-Martin S, Pike MC, Ross RK, Jones PA, Henderson B.Increased cell division as a cause of human cancer.Cancer Res1990;50:7415–7421. |
|43.||Branzei D, Foiani M.Regulation of DNA repair throughout the cell cycle.Nat Rev Mol Cell Biol2008;9:297–308. |
|44.||Delhommeau F, Dupont S, Tonetti C, Masse A, Godin I, Le Couedic J, et al..Evidence that the JAK2 (V617F) mutation occurs in a lympho-myeloid progenitor in polycythemia vera and idiopathic myelofibrosis.Blood2007;109:71–77. |
|45.||Vannuchi A, Antonioli E, Guglielmelli P, Longo G, Pancrazzi A, Ponziani V, et al..Prospective identification of high-risk polycythemia vera patients based on JAK2(V617F) allele burden.Leukemia2007;21:1952–1959. |
|46.||Vannuchi AM, Antonioli E, Guglielmelli P, Rambaldi A, Barosi G, Marchioli R, et al..Clinical profile of homozygous JAK2 V617F mutation in patients with polycythemia vera or essential thrombocythemia.Blood2007;110:840–846. |
|47.||Jones A, Silver R, Waghorn K, Curtis C, Kreil S, Zoi K, et al..Minimal molecular response in polycythemia vera patients treated with imatinib or interferon alpha.Blood2006;107:3339–3341. |
|48.||Ben Yehuda A, Globerson A, Krichevsky S, Bar On H, Kidron M, Friedlander Y, et al..Ageing and the mismatch repair system.Mech Ageing Dev2000;121:173–179. |
[Table 1], [Table 2], [Table 3]