Splanchnic vein thrombosis (SVT), a complication of myeloproliferative disorders (MPDs), sometimes reveals polycythemia vera (PV) or essential thrombocythemia (ET).1  However, polycythemia and thrombocytosis may be masked by hemodilution and/or hypersplenism in SVT patients, making the diagnosis of MPD difficult. In this retrospective study, SVT was diagnosed by Doppler ultrasound and/or multidetector spiral computed tomography for 45 patients with normal or low hematocrit and platelet counts. These patients were examined for the presence of 2 clonal markers specific for MPDs: the JAK2-V617F mutation2  and the capacity to form endogenous erythroid colonies (EECs) and/or endogenous megakaryocytic colonies (EMCs), observed in both mutated and nonmutated MPDs (Table 1).3-5  Levels of expression of JAK2 wild-type (WT) or mutated (V617F) were determined in blood granulocytes (37 patients) and/or in May-Grünwald-Giemsa–stained myeloid colonies (11 patients for whom granulocytes were not available) using allele-specific quantitative polymerase chain reactions (PCRs) adapted to complementary (granulocytes) or genomic (colonies) DNA.5,6  Formation of EECs/EMCs by bone marrow (BM) progenitors in collagen gels, serum erythropoietin, and granulocyte PRV-1 levels were also analyzed.3-5,7 

Table 1.

Comparison of SVT patients with and without latent MPD and the JAK2-V617F mutation



All patients

Patients with latent MPD

No evidence of MPD
Latent MPD
JAK2-V617F negative
JAK2-V617F positive
No. patients   28   17   3   14  
Site of thrombosis, hepatic veins/portal venous system   4/24   2/15   0/3   2/12  
Sex, male/female   15/13   9/8   3/0   6/8  
Age, y   51 ± 16   40 ± 11   40 ± 18   42 ± 11  
JAK2-V617F, copies/100 ABL   0   86 ± 91   0   110 ± 89  
JAK2-WT, copies/100 ABL   774 ± 889   491 ± 476   222 ± 155   564 ± 511  
Hematocrit   0.380 ± 0.57   0.396 ± 0.53   0.383 ± 0.42   0.399 ± 0.55  
Hemoglobin level, g/L   127 ± 20   130 ± 15   131 ± 16   129 ± 16  
Platelet count, × 109/L   239 ± 100   251 ± 123   237 ± 122   255 ± 128  
Leukocyte count, × 109/L   8.0 ± 5.4   7.8 ± 5.0   7.2 ± 2.9   7.9 ± 5.4  
PRV-1/ABL, CT ratio   0.962 ± 0.084   0.875 ± 0.120*  1.032 ± 0.024   0.843 ± 0.106 
EPO level, IU/L   24.5 ± 31.5   9.0 ± 8.9   6.7 ± 0.4   9.6 ± 9.9  
EECs, patients positive, %   0   12   0   14  
EMCs, patients positive, %   0   87   100   85  
Histology in favor of MPD, no./no. total   0/10   11/12   2/2   9/10  
Inherited thrombophilia, no./no. total  5/19   3/9  0/2   3/9  
    Protein C deficiency   2   0   0   0  
    Protein S deficiency   1   3   0   3  
    Heterozygous factor V Leiden mutation   1   1   0   1  
    Heterozygous factor II G20210A   1   0   0   0  
Acquired thrombophilia  8   0   0   0  
    Lupus anticoagulant   2   0   0   0  
    Pregnancy   1   0   0   0  
    Hepatocarcinoma   1   0   0   0  
    Cirrhosis   2   0   0   0  
    Severe acute pancreatitis   1   0   0   0  
    Abdominal sepsis
 
1
 
0
 
0
 
0
 


All patients

Patients with latent MPD

No evidence of MPD
Latent MPD
JAK2-V617F negative
JAK2-V617F positive
No. patients   28   17   3   14  
Site of thrombosis, hepatic veins/portal venous system   4/24   2/15   0/3   2/12  
Sex, male/female   15/13   9/8   3/0   6/8  
Age, y   51 ± 16   40 ± 11   40 ± 18   42 ± 11  
JAK2-V617F, copies/100 ABL   0   86 ± 91   0   110 ± 89  
JAK2-WT, copies/100 ABL   774 ± 889   491 ± 476   222 ± 155   564 ± 511  
Hematocrit   0.380 ± 0.57   0.396 ± 0.53   0.383 ± 0.42   0.399 ± 0.55  
Hemoglobin level, g/L   127 ± 20   130 ± 15   131 ± 16   129 ± 16  
Platelet count, × 109/L   239 ± 100   251 ± 123   237 ± 122   255 ± 128  
Leukocyte count, × 109/L   8.0 ± 5.4   7.8 ± 5.0   7.2 ± 2.9   7.9 ± 5.4  
PRV-1/ABL, CT ratio   0.962 ± 0.084   0.875 ± 0.120*  1.032 ± 0.024   0.843 ± 0.106 
EPO level, IU/L   24.5 ± 31.5   9.0 ± 8.9   6.7 ± 0.4   9.6 ± 9.9  
EECs, patients positive, %   0   12   0   14  
EMCs, patients positive, %   0   87   100   85  
Histology in favor of MPD, no./no. total   0/10   11/12   2/2   9/10  
Inherited thrombophilia, no./no. total  5/19   3/9  0/2   3/9  
    Protein C deficiency   2   0   0   0  
    Protein S deficiency   1   3   0   3  
    Heterozygous factor V Leiden mutation   1   1   0   1  
    Heterozygous factor II G20210A   1   0   0   0  
Acquired thrombophilia  8   0   0   0  
    Lupus anticoagulant   2   0   0   0  
    Pregnancy   1   0   0   0  
    Hepatocarcinoma   1   0   0   0  
    Cirrhosis   2   0   0   0  
    Severe acute pancreatitis   1   0   0   0  
    Abdominal sepsis
 
1
 
0
 
0
 
0
 

Patients with no evidence of MPD are patients who did not carry the JAK2-V617F mutation and did not form EMC/EEC in vitro. Patients with latent MPD are patients who carried the JAK2-V617F mutation and/or formed EMCs/EECs in vitro. Unless stated otherwise, results were expressed as means ± SD.

CT indicates cycle threshold.

*

P = .028 compared with patients with no evidence of MPD.

P = .009 compared with patients negative for JAK2-V617F, Mann-Whitney rank sum test.

One patient had 2 causes of inherited thrombophilia.

JAK2-V617F was detected in granulocytes from 12 of 37 SVT patients and in myeloid colonies from 2 of 8 additional patients; EMCs were observed for 14 patients (2 also formed EECs; Figure 1). Altogether, 14 (31%) of 45 SVT patients carried the JAK2-V617F mutation and 17 (38%) of 45 were positive for JAK2-V617F and/or EMCs/EECs, and were therefore considered to be suffering latent MPD. Two patients without EMCs were positive for JAK2-V617F, and 3 patients negative for JAK2-V617F had EMCs. BM histology, which was not always performed during anticoagulant treatment, was available for 22 patients and in favor of MPDs8  (2 PV, 9 ET) for 11 of 12 patients with latent MPDs (JAK2-V617F and/or EMCs/EECs); 1 of 2 patients with only EMCs rapidly developed typical ET. Histologic features of MPD were not observed in 10 of 10 SVT patients negative for both EMCs/EECs and the JAK2 mutation (Figure 1). In addition, a low serum erythropoietin level (< 3.3 IU/L) was found only in MPD patients,7  and 7 of 9 patients with high PRV-1 levels carried the JAK2-V617F mutation, an expected finding since PRV-1 expression correlates with JAK2 activation.9  Consistent with our previous observation of JAK2 overexpression in reactive states,5  the quantification of total JAK2 transcripts revealed that granulocytes of SVT patients expressed high levels of JAK2 whether they suffer from MPDs or not (Table 1). Importantly, the proportion of mutated JAK2 was typically low (median, 18%; range, 2%-51%), close to the values found in ET.5  Among the 3 patients with more than 40% JAK2-V617F, 1 had EECs and the other 2 had low erythropoietin levels; 2 later developed PV. Finally, acquired thrombophilia was found in 8 of 28 patients without MPDs but never in MPD patients; association with inherited thrombophilia was equally frequent in both groups of patients (Table 1; Figure 1).

Figure 1.

Patient characteristics. Results of the EMC, EEC, PRV-1, and erythropoietin (EPO) assays and bone marrow histology are shown according to the percentage of total JAK2 represented by JAK2-V617F; inherited and acquired thrombophilia are also shown. ▪ indicates positive EMC and EEC assays, increased PRV-1 expression (defined as a PRV-1 cycle threshold [CT]/ABL CT ratio < 0.850), low serum EPO level (< 3.3 IU/L), presence of inherited or acquired thrombophilia, and bone marrow histology in favor of PV (panmyelosis; increased cellularity predominating on the erythroid lineage; megakaryocytes in clusters, pleiomorphic, with deeply lobulated nuclei) or in favor of ET (increased cellularity predominating on the megakaryocytic lineage; megakaryocytes in loose clusters or dispersed, large to giant, with abundant, mature cytoplasm and hyperlobulated nuclei, lacking dysplastic features; absence of fibrosis).8  □ indicates negative EMC and EEC assays, normal PRV-1 expression, normal or high serum EPO level, absence of inherited or acquired thrombophilia, and normal bone marrow histology results. Empty spaces mean that the assay was not performed or that the results were not available.

Figure 1.

Patient characteristics. Results of the EMC, EEC, PRV-1, and erythropoietin (EPO) assays and bone marrow histology are shown according to the percentage of total JAK2 represented by JAK2-V617F; inherited and acquired thrombophilia are also shown. ▪ indicates positive EMC and EEC assays, increased PRV-1 expression (defined as a PRV-1 cycle threshold [CT]/ABL CT ratio < 0.850), low serum EPO level (< 3.3 IU/L), presence of inherited or acquired thrombophilia, and bone marrow histology in favor of PV (panmyelosis; increased cellularity predominating on the erythroid lineage; megakaryocytes in clusters, pleiomorphic, with deeply lobulated nuclei) or in favor of ET (increased cellularity predominating on the megakaryocytic lineage; megakaryocytes in loose clusters or dispersed, large to giant, with abundant, mature cytoplasm and hyperlobulated nuclei, lacking dysplastic features; absence of fibrosis).8  □ indicates negative EMC and EEC assays, normal PRV-1 expression, normal or high serum EPO level, absence of inherited or acquired thrombophilia, and normal bone marrow histology results. Empty spaces mean that the assay was not performed or that the results were not available.

In conclusion, detection of JAK2-V617F and EECs/EMCs, which can be performed during anticoagulant treatment, diagnosed a latent MPD, most often ET, in 38% of patients with SVT and normal/low blood cell counts. We propose that at least the presence of JAK2-V617F in blood granulocytes should be systematically investigated in such situations.

M.B. and E.L. contributed equally to this study.

Supported by grants from the Programme Hospitalier de Recherche Clinique (PHRC) of the Région Bourgogne and the Délégation Régionale à la Recherche Clinique (DRRC) of the Région Pays de Loire and the CHU de Nantes.

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