Pathologie Biologie 55 (2007) 92104

http://france.elsevier.com/direct/PATBIO/

Current diagnostic criteria for the chronic myeloproliferative

disorders (MPD) essential thrombocythemia (ET),
polycythemia vera (PV) and chronic idiopathic myelofibrosis (CIMF)

Critres diagnostiques actuels des syndromes myloprolifratifs (SMP),

thrombocytmie essentielle (TE), polyglobulie de Vaquez (PV)
et mylofibrose idiopathique (MFI)
J.J. Michielsa,d,*, Z. Bernemaa, D. Van Bockstaeleb, H. De Raevec, W. Schroyensa
a

Department of Hematology, University Hospital Antwerp Wilrijkstraat 10, 2650 Edegem/Antwerp, Belgium
b
Department of Molecular Genetics, University Hospital Antwerp, Antwerp, Belgium
c
Department of Pathology University Hospital Antwerp, Antwerp, Belgium
d
Goodheart Institute, Hematology, Hemostasis and Thrombosis Research Center,
Rotterdam, MPD Center Europe, Erasmus Tower, Veenmos 13, 3069 AT Rotterdam, The Netherlands
Available online 21 August 2006

Abstract
The clinical criteria for the diagnosis of essential thrombocythemia (ET) according to the polycythemia vera study group (PVSG) do not
distinguish between ET and thrombocythemia associated with early stage PV and prefibrotic chronic idiopathic myelofibrosis (CIMF). The
clinical criteria of the PVSG for the diagnosis of polycythemia vera (PV) only detects advanced stage of PV with increased red cell mass. The
bone marrow criteria of the World Health Organization (WHO) are defined by pathologists to explicitly define the pathological criteria for the
diagnostic differentiation of ET, PV, and prefibrotic and fibrotic CIMF. As the clinical PVSG and the pathological WHO criteria show significant
shortcomings, an updated set of European Clinical and Pathological (ECP) criteria combined with currently available biological and molecular
markers are proposed to much better distinct true ET from early PV mimicking ET, to distinguish ET from thrombocythemia associated with
prefibrotic CIMF, and to define the various clinical and pathological stages of PV and CIMF that has important therapeutic and prognostic
implications. Comparing the finding of clustered giant abnormal megakaryocytes in a representative bone marrow as a diagnostic clue to
MPD, the sensitivity for the diagnosis of MPD associated with splanchnic vein thrombosis was 63% for increased red cell mass, 52% for low
serum EPO level, 72% for EEC, and 74% for splenomegaly indicating the superiority of bone marrow histopathology to detect masked early and
overt MPD in this setting. The majority of PV and about half of the ET patients have spontaneous EEC, low serum EPO levels and PRV-1 overexpression and are JAK2 V617F positive. The positive predictive value for the diagnosis of PV of spontaneous growth of endogenous erythroid
colonies (EEC) of peripheral blood (PB) and bone marrow (BM) cells is about 8085% when either PB or BM EEC assays, and up to 94% when
BM and PB EEC assays were performed. The diagnostic impact of low serum EPO levels (ELISA assay) in a large study of 186 patients below
the normal range (< 3.3 IU/l) had a sensitivity specificity and positive predictive value of 87%, 97% and 97.8%, respectively, for the diagnosis of
PV. There is a significant overlap of serum EPO levels in PV versus control and controls versus SE. The specificity of a JAK2 V617F PCR test
for the diagnosis of MPD is high (near 100%), but only half of ET and MF (50%) and the majority of PV (up to 97%) are JAK2 V617F positive.
The use of biological markers including JAK2 V617 PCR test, serum EPO, PRV-1, EEC, leukocyte alkaline phosphatase score and peripheral

Lecture International Symposium on The V616F JAK2 mutation: A major step forward in the pathogenesis and management of myeloproliferative disorders,
November 18, 2005, Hopital Avicenne and Paris 13 University.
* Corresponding author.
E-mail address: postbus@goodheartcenter.demon.nl (J.J. Michiels).
0369-8114/$ - see front matter 2006 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.patbio.2006.06.002

J.J. Michiels et al. / Pathologie Biologie 55 (2007) 92104

93

blood parameters combined with bone marrow histopathology has a high sensitivity and specificity (almost 100%) to diagnose the early and overt
stages of ET, PV and CIMF in JAK2 V617F positive and negative MPDs.
2006 Elsevier Masson SAS. All rights reserved.
Rsum
Les critres cliniques de diagnostic de la thrombocytmie essentielle (TE) dvelopps par le PVSG ne permettent pas de distinguer les TE des
formes dbutantes avec thrombocytose de PV et de MFI. Ces critres ne permettent par ailleurs que le diagnostic de formes patentes de PV, avec
une claire augmentation du volume globulaire. Les critres OMS de diagnostic des SMP incluent la biopsie mdullaire, avec des critres dfinis
par les pathologistes pour clairement diffrencier TE, PV, MFI au stade prfibrotique ou fibrotique. Nous proposons une mise jour des critres
diagnostiques utilisant les marqueurs biologiques et molculaires actuellement disponibles, dans le but de distinguer des TE pures les formes
dbutantes de PV et de MFI mimant les TE, et de dfinir les stades cliniques et anatomopathologiques de PV et MFI, ce qui a dimportantes
consquences pronostiques et thrapeutiques. Sont notamment discuts, la valeur des donnes morphologiques sur la biopsie de moelle osseuse,
du dosage drythropotine, de lexistence de colonies rythrodes ou mgacaryocytaires spontanes, de lhyperexpression du gne PRV-1, et de
la mutation V617F de JAK2. Lutilisation de ces marqueurs en association avec les donnes morphologiques de la moelle osseuse permet de
diagnostiquer avec une haute sensibilit et spcificit (proches de 100 %) les stades patents mais aussi prcoces de TE, PV et MFI, quils soient
V617F JAK2 positifs ou ngatifs.
2006 Elsevier Masson SAS. All rights reserved.
Keywords: Myeloproliferative disorders; Essential thrombocythemia; Polycythemia vera; Myeloid metaplasia; Myelofibrosis; Erythropoietin; Endogenous erythroid
colony assay; JAK2 V617F mutation; Bone marrow pathology
Mots cls : Thrombocytmie ; Polyglobulie ; JAK2 ; Biopsie mdulllaire

1. Introduction
In 1950, William Dameshek presented an original view on
the physiopathology and course of polycythemia vera (PV) [1].
He described PV as a chronic disorder of the bone marrow
characterized by excessive production of blood cells by the
marrow elements i.e. the nucleated red cells the granulocytes
and the megakaryocytes. Not only is PV a chronic disorder
without any evidence of invasiveness but it is a total marrow
disorder in which erythrocytosis, leukocytosis and thrombocytosis are all simultaneously present [1]. In PV, all stops to
blood production in the bone marrow seem to have been pulled
out. The marrow is crowded with great numbers of nucleated
red cells and granulocytes in all stages of maturation with
marked hyperplasia and clustering of enlarged mature megakaryocytes. Some cases however show only a moderate elevation of erythrocytes with an extreme degree of thrombocytosis,
while in others the leukocyte counts may be at or close to leukemic levels, with only slight increase in red cells or platelets.
As to the etiology of trilinear myeloproliferation in PV, Dameshek proposed two highly speculative possibilities one, the presence of excessive bone marrow stimulation by an unknown
factor or factors and two, a lack or a diminution in the normal
inhibitory factor or factors [1]. This original view and hypothesis of Dameshek is recently confirmed by the discovery of the
JAK2 V617F mutation by James, Ugo, Casadevall and Vainchenker in Paris, France [2] demonstrating that the V617F mutation induces a loss of inhibitory activity of the JH2 pseudokinase part on the JH1 kinase part of JAK2 leading to enhanced
activity of the normal JH1 kinase activity of JAK2, which
makes the mutated hematopoietic stem cells hypersensitive to
hematopoietic growth factors TPO EPO, IGF1, SCF and GCSF
resulting in trilinear myeloproliferation.

2. The concept of PV as a trilinear MPD

Wasserman proposed in 1954 a hypothetical concept for the
course of PV[3,4]. Accordingly PV is characterized by initial
erythrocytosis (erythrocythemia), which is usually accompanied by leukocytosis of normal mature granulocytes (leukocythemia) with increased leukocyte alkaline phosphatase
score, by thrombocytosis (thrombocythemia), splenomegaly,
and plethora. Wasserman distinguished at least five subsequent
stages in the natural history of PV (Fig. 1) [3,4]:
stage 1). Pure erythrocythemia is featured by increased
hemoglobin, hematocrit and red cell mass with normal leukocytes, thrombocytes and spleen size, which is labeled as
idiopathic erythrocytosis by Pearson and Whetherley-Mein
[5] and Najean et al. [6]. This category of early PV may
comprise about 2030% of the cases at time of presentation;
stage 2). The polycythemic stage of PV is featured by
thrombocythemia, erythrocythemia and no or slight myeloid
metaplasia, leukocytosis and/or splenomegaly (Fig. 1);
stage 3). Myeloid metaplasia in PV patients presents with
no or different grades of reticulin and collagen fibrosis in
the bone marrow and progressive splenomegaly during
long-term follow in about one third of the cases;
stage 4). The polycythemic stage with various degrees myelofibrosis and splenomegaly following PV may elapse 5
25 years before a period of normal red cell values socalled spontaneous remission of PV occurs. This stage
must be considered as the beginning of spent phase PV
and may last a few to several years. At this point the spleen
is frequently large and very firm to palpation, the liver is
enlarged to a moderately degree in most patients, thrombocythemia is frequent and may be pronounced with bizarre
and giant platelets, and white cells are usually increased

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Based on extended clinical experience and review of the

literature, Glaser and Walker [7] concluded that the transition
of PV to myelofibrotic myeloid metaplasia occurs, but whether
post-PV is the same or distinct from agnogenic myeloid metaplasia (AMM) or chronic idiopathic myelofibrosis (CIMF) still
remains an open question and has never been solved in a prospective clinicopathological study, which nowadays becomes
possible by the use of new molecular and biological markers
including PRV-1 expression and JAK2 V617F mutation [2].
3. Clinical, laboratory and pathological features
of the MPDs

Fig. 1. The evolution and dynamics of the disease process in polycythemia vera
(PV) according to the concept Wasserman, who defined PV as a trilinear
myeloproliferative disorder with various degrees of erythrocythemia, thrombocythemia, leukocythemia and prefibrotic myeloid metaplasia as initial stages
followed by spent phase PV and dry tap myelofibrosis after long-term followup in about one third of the cases [3,4].

with granulocytic leukocytosis (leukocythemia) accompanied by a small percentage of immature forms;

stage 5). Post-PV myeloid metaplasia shows various degrees
of leuko-erythroblastosis of the peripheral blood and may
progress to extreme myelofibrosis with a dry tap on aspiration and massive splenomegaly (Fig. 1). At this end-stage
histopathology of bone marrow biopsy shows a similar picture and can not been differentiated from agnogenic myeloid
metaplasia with no previous history of PV [1,2].

In the 1980s and 1990s the German pathologists Burkhardt

et a1 [8], Georgii et al. [912] and Thiele et al. [1318] have
described typical histopathological features from bone marrow
biopsy material for the diagnosis and classification of each of
the 3 different Ph-negative MPDs essential thrombocythemia
(ET), PV and AMM. Georgii et al. [9,10] and Thiele et al.
[13,14] drew attention to an authentic chronic megakaryocytic
granulocytic myeloproliferation (CMGM) as a separate pathological entity among the MPD, distinct from ET and distinct
from chronic myeloid leukemia (CML) or myelodysplastic
syndrome (MDS). This condition has been described in 1996
as Philadelphia-chromosome negative chronic megakaryocytic
granulocytic myeloproliferation (CMGM) by Georgii et al. [11,
12] and as prefibrotic CIMF by Thiele et al. [18]. Since the
early 1980s Michiels and Juvonen [1921] combined clinical
and pathological features by including bone marrow histopathology as a pathognomonic, diagnostic clue to each of the
MPDs. In 1997, the thrombocythemia vera study Group
(TVSG) extended the clinical criteria of the polycythemia
vera study Group (PVSG) for the diagnoses of ET and PV by
including bone marrow histopathology as a diagnostic clue and
specific feature of ET, PV (Tables 1 and 5A) [21]. Thiele et al.
[22,23] improved the TVSG criteria for ET, PV and added the
Cologne criteria for prefibrotic, early fibrotic and overt fibrotic
stages of CIMF (Table 2). These clinical and pathological criteria were diagnostic for MPD [2023] and inspired the World
Health Organization (WHO) [24] to clearly define the pathological bone marrow criteria for the differentiation between true

Table 1
The World Health Organization (WHO) [24] and the European Clinical and Pathological (ECP) [25] criteria for the diagnosis of acquired or congenital essential
thrombocythemia: ET
Clinical criteria
Persistent increase of platelet count
ECP: > 400 109/l
WHO: > 600 109/l
ECP
Presence of large or giant platelets in peripheral blood smear
ECP
Absence of any underlying disorder for reactive thrombocytosis
WHO and ECP
No peripheral blood, bone marrow and cytogenetic evidence of
PV, CML, CIMF, MDS or reactive thrombocytosis
ECP
Absence of any cytogenetic abnormality

Pathological criteria
WHO and ECP
Increase of dispersed or loosely clustered, predominantly enlarged mature megakaryocytes with
hyperlobulated nuclei and mature cytoplasm, normal cellularity, no or borderline increase of
reticulin
A typical ET picture excludes PV, AMM, CML, MDS, and reactive thrombocytosis
ECP
No proliferation or immaturity of granulo- or erythropoiesis
Molecular biology: ECP
Clonality studies: polyclonal or monoclonal
Acquired: JAK2 V617F positive or negative
Congenital: polyclonal and JAK2 V617F negative , caused by gain of function mutation of
TPO, cMPL genes or of unknown etiology

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95

Table 2
The World Health Organization (WHO) [24] and the updated European Clinical and Pathological (ECP) [25] Criteria for the diagnosis of prefibrotic and early
fibrotic agnogenic myloid metaplasia (AMM)
Clinical criteria, ECP
No preceding or allied other subtype of myeloproliferative
disorders CML or MDS.Main presenting feature of prefibrotic
CIMF is thrombocythemia and slight splenomegaly, no dry tap
on bone marrow aspiration and diagnosed as ET according to
the PVSG.
Clinical and laboratory features
Normal hemoglobin or slight anemia, grade I: hemoglobin > 12
g/dl
Slight or moderate splenomegaly on ultrasound scan or CT
Thrombocytosis, platelets in excess of 400, 600 or even
1000 109/l
No leuko-erythroblastosis
No tear drop erythrocytes

Pathological criteria, WHO, ECP

Chronic megakaryocytic and granulocytic myeloproliferation (CMGM) [912,20] and no or
relative reduction of erythroid precursors. Abnormal clustering and increase in atypical giant to
medium sized megakaryocytes containing bulbous (cloud-like) hypolobulated nuclei and
definitive maturation defects
European consensus on grading of myelofibrosis (MF) [80]
MF 0: Prefibrotic CIMF: focal fine reticulin with no intersections (cross-over) and only rare
course reticulin fibers
MF 1: Early fibrotic CIMF: loose network of reticulin with intersections, especially in
peripheral areas, no collagenization
Note that MF is not a disease but secondary to AMM
Molecular biology:
JAK2 V617F positive or negative
Screen for cytogenetic abnormalities

ET, classical PV, and prefibrotic and fibrotic stages of CIMF.

The WHO criteria are essentially drawn by international
experts in bone marrow morphology but not clinicians. To
overcome the shortcomings of the clinical PVSG and the
pathological WHO [24] criteria, Michiels and Thiele [25,26]
have defined a set of European Clinical and Pathological
(ECP) criteria, which by including the available biological
and molecular markers (Tables 1, 2, 5B, 7, 8) will much better
allow to diagnose and classify each of early and overt stages of
the MPDs ET, PV and CIMF [25,26].
4. WHO pathological criteria for the diagnosis of ET, PV
and CIMF
A typical bone marrow picture according to the WHO [24]
for true ET affects mainly of megakaryocytic cell lineage,
shows increased numbers of loosely clustered enlarged, mature
megakaryocytes with hyperploid staghorn-like nuclei together
with normal cellularity, normal erythropioesis, and normal
granulopoiesis, no increase of reticulin fibrosis, and there is
no peripheral blood, bone marrow and cytogenetic evidence
of typical and atypical chronic myeloid leukemias (CML),
PV, myelodysplastic syndrome (MDS) or reactive thrombocytosis (Table 1) [2026]. A typical bone marrow picture for true
ET excludes any other MPD, as well as typical and atypical
CML, MDS and reactive thrombocytosis.
A typical bone marrow picture according to the WHO [24]
for a prefibrotic form of CIMF is characterized by a prominent
granulocytic and megakaryocytic myeloproliferation (CMGM)
[11,12,20] and no or borderline increase in reticulin (CIMF-0,
Table 2). The prevalence of the (left-shifted) neutrophil and
megakaryocytic lineage is associated with reduction and
maturation arrest of erythroid precursors. Most conspicuous,
however is the fact that the megakaryopoiesis is characterized
not only by a disturbance of bone marrow histotopography
(loose to dense clustering and translocation to the endosteal
borders), but also by striking abnormalities of maturation.
The immature megakaryocytes in prefibrotic CIMF consist of
variations in size including giant forms and deviations of the

nuclear-cytoplasmic ratio accompanied by bulbous and hyperchromic cloud-like nuclei, which are never seen in ET and PV.
Thiele et al. [18,2736] demonstrated that prefibrotic and early
fibrotic CIMF usually presents with thrombocythemia (false
ET), which has to be distinguished from true ET because clinical features, natural history and prognosis significantly differ
(Table 2). The prefibrotic stage of CIMF is not only associated
with pronounced thrombocythemia, but also show no leukoerythroblastic blood picture, normal or increased LAF-score
and no or minimal splenomegaly (Table 2) [18,2736], and
therefore diagnosed as ET according to the PVSG [37,38] criteria (Tables 14).
A typical picture in the bone marrow according to the WHO
[24] pathognomonic and diagnostic for PV is featured by
increase of clustered enlarged mature megakaryocytes comparable to ET, and a moderate to marked increased cellularity,
erythropoiesis and granulopoiesis (i.e. panmyelosis) (Table 5A,
5B) [2026]. Bone marrow features in polycythemic stage of
PV show a hypercellular bone marrow of prominent erythroid
precursor cells and neutrophil granulopoiesis in addition to
megakaryocytes proliferation with loose arrangements of clustered megakaryocytes. Megakaryocytes do not only show different sizes, but fail to exhibit significant maturation defects.
The megakaryocytes in PV usually have a rather pleiomorphic
appearance with wide ranges of sizes including small and giant
forms. A typical PV picture of the bone marrow is seen in
Table 3
PVSG [38] criteria for the diagnosis of ET
Platelet count > 600 x 109/l: 1986
No known cause of Reactive Thrombocytosis
Normal hemoglobin and red cell mass to exclude overt PV
Stainable iron bone marrow to exclude PV
No features of MDS in bone marrow smear and biopsy
Absence of Ph1+ chromosome (brc/abl) to exclude CML
Collagen fibrosis of bone marrow is allowedup to <1/3 of bone marrow
biopsy area,without splenomegaly or leukoerythroblastic blood picture and
therefore includes thrombocythemia associated with prefibrotic and early
fibrotic myeoid metaplsia (MM)
A diagnosis of exclusion reactive thrombocytosis, overt PV, MDS, CML and
inclusion of prefibrotic and early fibrotic stages of myeloid metaplasia (MM).

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Table 4
ET according to PVSG [38] compared to WHO [24] and ECP [25] criteria
ET PVSG
Includes
Incidence
Serum EPO

Hereditary ET

True ET
2030
Normal

Earlyl PV
Mimicking ET
2030
Decreased

Prefibrotic CIMF
False ET
4060
Normal

< 0.001
Normal

Platelet

> 400 ECP

> 600 WHO
N
N
ET picture

> 400 ECP

> 600 ECP
N/
N/
PV picture

> 400 ECP

> 600 WHO
N/
N/
MMM picture
abnormal

Monoclonal

Erythrocytes
Hematocrit
Bone marrow:
Megakaryocytes
Splenomegaly
JAK2 V617F
EEC
PVR-1
Clonality

N
N
ET picture
Normal large / giant and mature

Neg:

Neg:

Not applicable

polyclonal
polyclonal monoclonal

++
++
++
Monoclonal

Table 5A
Extension of the PVSG [39] criteria of polycythemia vera (PV) by including bone marrow histopathology as a diagnostic clue to early and overt stages PV [21]
The Rotterdam Criteria of Polycytemia Vera Proposed by the thrombocithemia Vera Study group (TVSG)
Raised red cell mass
B1
Thrombocytosis
males > 36mL/Kg
Platelet count > 400 109/L
A2
Absence of any cause of secondary erythrocytosis by clinical and
B2
Granulocytes > 10 109/L and/or raised neutrophil alkaline
phosphatase score of > 100 in the absence of fever or infection
laboratory investigations
A3
Histopathologie of bone marrow biopsy increase of:
B3
Splenomegaly on palpation or isotope/ultrasound scan
a. cellularity, panmyelosis
b. enlarged megakaryocytes with hyperploid nuclei;
c. reticulin fibers (optional)
B4
Erythroid colony formation in absence of EPO: spontaneous EEC
A1 + A2 + A3 is consistent with early stage PV (so-called "idiopathic erythrocytosis")
A1 + A2+ A3 + any one from category B establishes overt PV
A3 + B1 is consistent with essential thrombocythemia
A3 + B3 and/or B4 is consitent with a primary myeloproliferative disorder
A1

classical PV according to the PVSG [39], in erythrocythemia

or idiopathic erythrocytosis, in early or latent PV and in
masked PV (Fig. 2 and Tables 6 and 7) [25,26,4044]. In congenital polycythemia (CP) and secondary erythrocytosis (SE),
in which increased erythropoiesis is present, the number, size,
morphology and distribution of megakaryocytes in bone marrow smears and biopsies remain normal [2023,40,41].
5. Current clinical and pathological criteria
for the diagnoses of ET PV and CIMF
Since 1975 the minimum criterion of the Polycythemia Vera
Study Group (PVSG) for the diagnosis of ET was 1000 109/l
(1 million) [37]. In 1985, we demonstrated that the majority of
symptomatic ET patients diagnosed by increase and clustering
of enlarged megakaryocytes in bone marrow biopsies, had platelet counts between 400 and 1000 109/l (below 1 million).
This prompted the PVSG to lower the platelet count for the
diagnosis of ET to the arbitrary minimum of 600 (Table 3)
[38]. We could consistently demonstrate that the arbitrary
minimum of 600 109/l platelets according to the PVSG and
WHO overlooks early stage MPD at platelets between 400 and
600 with no or slight splenomegaly, with no or low serum
EPO, and typical MPD features in the bone marrow consistent

with early ET (Table 1), early PV mimicking ET (Tables 4,

5A, 5B and 6) or prefibrotic CIMF or unclassifiable MPD
(Tables 1 and 3).
The PVSG criteria for ET [38] is a diagnosis of exclusion of
reactive thrombocytosis, PV, MDS and Ph1+ CML but includes
by definition thrombocythemia associated with prefibrotic and
early fibrotic CIMF (Table 3) [1723]. The WHO [24] and the
ECP [25] criteria for the diagnoses of ET extend the PVSG
[34] criteria by including histopathology from bone marrow
biopsies as a positive criterion of MPD and as powerful tool
to differentiate between true ET, false ET and early PV
mimicking ET (Table 4). Comparing the WHO and ECP with
the PVSG criteria for the diagnosis of ET show that the PVSG
criteria fail to distinguish ET from early PV mimicking ET
[4044] and fail to distinguish ET from usually pronounced
thrombocythemia associated with prefibrotic CIMF (Tables 4)
[2325,3336]. In consideration of disease-related complications occurring at low platelet counts, the arbitrarily chosen
limit for platelet count (> 600 10/l) by the PVSG (Table 3)
[38] and WHO (Table 1) [24] has been reduced to 400 109/l
in the ECP [25] (Table 1) criteria for ET. The PVSG [38] criteria for ET, when compared to the WHO [24] and ECP [25]
criteria, also include early PV mimicking ET (Tables 4 and 6)
[4044].

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97

Table 5B
Diagnosis of PV according to the PVSG, WHO and ECP criteria
Criteria
A1
Overt
PV

PVSG [39]
Major criteria
Red cell mass: RCM
Male > 36 ml/kg
Female > 32 ml/kg

Latent PV

A2
A3
A4
A5
Criteria
B1
B2
B3

Normal arterial oxygen saturation > 92%

Splenomegaly on palpation

Minor criteria
Platelets > 400
Leukocytes > 12
Bone marrow biopsy has been disregarded by
the PVSG and included by the TVSG as a
diagnostic clue for early and overt PV [21]

B4

Raised LAP score

Diagnosis
A1 + A2 + A3
A1 + A2 + two from B
Manifest PV:
Increased RCM

WHO [24]
Major criteria
Red cell mass: RCM
> 25% above mean
normal value
or Hb > 18.5 g/dl in men, Hb > 16.5 g/dl in
women

Absence secondary erythrocytosis

Splenomegaly on palpation
Clonal evidence other than Ph1+ or BCR/ABL
Spontaneous EEC
Minor criteria
Platelets > 400
Leukocytes > 12
Bone marrow biopsy with typical PV picture
Increased cellularity with trilineage myeloproliferation and clustering of small to giant (pleiomorphic) megakaryocytes
Low serum EPO
Diagnosis
A1 + A2 + any other from A
A1 + A2 + two from B
Manifest PV:
Increased RCM

ECP [25]
Clinical criteria
Red cell mass optional
> 25% above mean normal value
or Hb > 18.5 g/dl in men
Hb > 16.5 g/dl in women
Red cell mass normal
and Ht < 0.51 in men
Ht < 0.48 in female
Absence of secondary erythrocytosis
Splenomegaly on CT or ultrasound (> 12 cm)
Clonal evidence other than Ph1+ or BCR/ABL
Spontaneous EEC
Clinical criteria
Platelets > 400 109/l
Leukocytes >12 109/l
Bone marrow biopsy with typical PV picture
Increased cellularity with trilineage myeloproliferation and clustering of small to giant (pleiomorphic) megakaryocytes
Low serum EPO
Diagnosis
B3 plus any other of the clinical criteria
Manifest PV:
Increased RCM
Early stage PV:
RCM Normal

Table 6
Clinical staging of Polycythemia Vera: therapeutic implications
Polycythemia Vera
Staging of PV as one distinct
disease
Incidence (%)
Hemoglobin g/dl
Serum EPO
Hematocrit
Red cell mass
Thrombocytes (x 109/l)
Leukocytes (x 109/l)
Spleen on echogram
Bone marrow:
JAK2 V617F
EEC
PRV-1 LAF score
Myelofibrosis (MF)

Evolution
ECP 1 Aspirin/phlebotomy

ECP 2 Aspirin/phlebototm

Manifestation
ECP 3 PVSG WHO A/P

initial PV mimicking ET
2025
N/

Erythrocythemic PV
2025

Thrombo/erythro/leukocythemic PV
4060

Male 0.43- < 0.51

Female 0.42- < 0.48
N
>400600
N
N15 cm
PV picture
+
+

Male > 0.51

Female > 0.48

< 400
N
N
PV picture
+
+

> 400
N
N15 cm
PV picture
+/++
+

0/1

> 1,000
> 15
> 15 cm
PV MF picture
++/LOH
+

1/2

The PVSG have used increased red cell mass proposed by

Dameshek in 1950 [1] as the main inclusion criterion for diagnosis of PV in the PVSG 01 study since 1969 [4], which
became a main diagnostic criterion for the diagnosis of PV
since 1975 (Tables 5A, 5B) [39]. Increased red cell mass is
not specific, because it includes patients with PV, congenital
polycythemias (CP), and secondary erythrocytosis (SE),
thereby introducing a main differential diagnostic problem
and the need of extensive laboratory investigations [45,46].

ECP4 PVSG WHO IFN/HU

The PVSG [39] postulated three major and four minor criteria
(Table 5B) to ensure that patients who entered the PVSG prospective trial indeed were suffering from PV and not from congenital polycythemia or secondary erythrocytosis [39].
Increased red cell mass corresponded with high hematocrit
values between 0.50 and 0.75 [39] (ECP stage 3 and 4 Table 6),
which according to the PVSG 01 study [39] was associated
increased platelet count in two third and with major thrombosis
in one third of more than 400 PV patients at time of diagnosis

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J.J. Michiels et al. / Pathologie Biologie 55 (2007) 92104

[4,39]. The thrombocythemia vera study group (TVSG,

Table 5A) [21] extended the PVSG criteria by including bone
marrow histopathology, which prompted pathologists to define
the WHO [24] and clinicians to define ECP [25] criteria for the
diagnosis of early or masked PV and overt and advanced
stages of PV (Tables 4 and 6).
According to the TVSG [21] criteria in Table 5A a typical
PV picture of the bone marrow is seen in 4 different categories
of MPD patients.
First, the combination of a typical PV picture, increased red
cell mass, high hematocrit and one of the B criteria (Table 5A)
is consistent with classical PV according the PVSG [39] and
WHO [24] and as advanced stage 3 and 4 PV when applying
the ECP [25] criteria (Table 6).
Second, a typical PV picture of the bone and increased red
cell mass, high hematocrit but with normal platelet count and
spleen size is consistent with erythrocythemia according to
Wasserman [3] (Fig. 1), diagnosed as idiopathic erythrocytosis
by the PVSG criteria [5,6], and diagnosed as erythrocythemic
PV according to WHO [24] and stage 2 ECP [25] when combined with serum EPO levels and EEC (Tables 5B and 6). In
contrast to the PVSG [39] criteria, both the WHO [24] and
ECP [25] criteria clearly differentiates between PV, CP and
SE without the need of red cell measurement in PV, and with
a clear indication for red cell mass measurement in CP and SE
[21,25,26].
Third, a typical PV picture of the bone marrow with a
hematocrit in the upper limit of normal but with increased platelet count is consistent with ET according to PVSG [39] and
WHO [24] but diagnosed as early stage 1 PV mimicking ET
according to the ECP [25] criteria (clinical case 2, Fig. 2 and
Tables 4, 5A, 5B and 6) [25]. Early stage 1 PV usually presents with microvascular disturbances at platelet count in
excess of 400 109l, increased LAF score, low serum EPO,
spontaneous EEC and no or slight splenomegaly (Tables 4
and 6) [26].
Fourth, the combination of a typical PV picture, normal red
cell mass, normal hemoglobin and hematocrit, normal platelet
but slowly progressive splenomegaly, granulocytosis or even

Fig. 2. The evolution and dynamics of the disease process in polycythemia vera
(PV) according to Thiele et al. [4044] indicating the sequential occurrence of
the early initial stage of PV mimicking ET, the overt polycythemic stage of
classical PV, and progression to post-PV myeloid metaplasia or leukemia as
terminal stages in about one third of the cases.

slight anemia is not consistent with either ET, PV but with

unclassifiable MPD or masked PV [21]. Such cases of unclassifiable MPD and masked PV are overlooked by clinicians and
may comprises about one quarter of patients with a typical PV
picture of the bone marrow. Such cases may present with
thrombotic complications including splanchnic vein thrombosis
[47,48], are described as masked PV [48,49], and frequently
show spontaneous EEC as the clue to the atypical presentation
of MPD. Masked ET or PV may progress to so-called classical CIMF without overt PV.
Using clusters of pleiomorphic giant megakaryocytes in
bone marrow biopsy as a reference standard for the diagnosis
MPD, a French study demonstrated that 46 out of 128 consecutive patients with splanchnic (hepatic, portal or mesenteric)
vein thrombosis had features of MPD [48]. These 46 MPD
patients with splanchnic vein thrombosis had hepatic vein
thrombosis (Budd Chiari syndrome) in 19 and portal or mesenteric vein thrombosis in 27 [48]. As compared to a positive
bone marrow finding of clustered giant megakaryocytes diagnostic for MPD, the sensitivity for the diagnosis of MPD associated with splanchnic vein thrombosis was 63% for increased
red cell mass, 52% for low serum EPO level, 72% for EEC,
and 74% for splenomegaly indicating the superiority of bone
marrow histopathology to detect masked early and overt stages
of MPD in this setting [48].
Thiele et al. [4044] confirmed the sequential occurrence of
early PV mimicking ET, overt and advanced stages of PV
(Fig. 2 and Table 6). The WHO [24] and ECP [25] criteria
clearly differentiate PV from CP, SP, and ET from reactive
thrombocytosis, initial PV and prefibrotic CIMF. The PVSG
[39] and the WHO [24] criteria used increased red cell as a
mandatory main criterion (Table 5B), which is a very crude
and therefore overlook by definition early PV mimicking true
ET (ECP stage 1 in Table 6) and the erythrocythemic phase
(ECP stage 2 in Table 6) of PV, formerly labeled as idiopathic
erythrocytosis [5,6]. Accumulating data from the literature
show that spontaneous endogenous erythroid colony formation
(EEC) [5153], low serum EPO [5460] and PRV-1 expression [6164] are the hall mark of PV. About 50% of ET
patients according to the PVSG are EEC positive [51,6569],
PRV-1 positive [6164], and may be associated with low
serum EPO [50,5759] indicating early PV mimicking ET
(Tables 6 and 7). The reports on EEC/PRV-1 positive ET diagnosed according to the PVSG [39] may have with low serum
EPO and therefore represent early ECP stage 1 of PV (Tables 6
and 7). The cohorts of early ECP stage 1 and 2 PV and the
overt ECP stage 3 PV patients are featured by spontaneous
EEC, positive PRV-1, low serum EPO levels and a typical
PV bone marrow picture (Fig. 2 and Tables 6 and 7) [5466].
Both the early (ECP stage 1 and 2) and overt (ECP stage 3) PV
patients are at high-risk for potential minor and major vascular
complications, because they present with elevated platelet
count mimicking true ET [65,66]. EEC/PRV-1 positive ET
patients according to PVSG criteria have a high risk of developing microvascular and major thrombotic complication as
compared to EEC/PRV-1 negative ET [65,66]. PRV-1-negative

J.J. Michiels et al. / Pathologie Biologie 55 (2007) 92104

99

Table 7
The updated European Clinical and Pathological (ECP) Criteria for the Diagnosis of Early, Overt and Advanced Stage Polycythemia Vera (PV)
Clinical (C) criteria suspected for PV
C 1. Classical PV: Red Cell Mass optional Hemoglobin > 18.5/> 16.5 g/dl
male/female Hematocrit (Ht) > 51/> 48% male/female
C 2. Early or latent stage PV
Hematocrit (Ht): 0.450.51 male and 0.430.48 female
C 3. Low plasma Epo level (ELISA)
C 4. Persistent increase of platelet count: grade I: 4001500, grade II: > 1500.
C 5. Splenomegaly on palpation or on ultrasound echogram (>12 cm length in
diameter).
C 6. Granulocytes > 10 109/l or Leukocytes > 12 109/l and/or raised LAPscore or increased PRV-1 expression in the absence of fever or infection.
C 7. Platelet-mediated microvascular ischemic, thrombotic complications
C 8. Typical PV signs and symptoms of hypervolumemia
C 9. Itching, fatigue, upper abdominal complaints
C 10. Absence of any cause of secondary erythrocytosis.

ET comprises a pathophysiologically distinct subgroup of true

ET patients with no features of early PV, who are at lower risk
for the development of thrombotic complications and for emergence of PV [66]. Early or initial PV (or forme fruste of PV)
according to updated ECP criteria in Table 7 is typically featured by a PV picture in the bone marrow, positive results for
EEC and PRV-1, and/or low serum Epo levels, and a high
thrombotic risk, which is related to increase of hypersensitive
platelet counts (thrombocythemia) and slight increase of hematocrit up to 0.50, and therefore candidates for low dose aspirin
and phlebotomy (Table 6).
Spontaneous growth of endogenous erythroid colonies
(EEC) and serum erythropoietine (EPO) levels are important,
but have insufficient diagnostic specificity and sensitivity as
isolated parameters to differentiate between PV and SE. The
reliability of peripheral blood (PB) and bone marrow (BM)
EEC was investigated in a multicenter study including 140
patients (80 PV according to the PVSG, 54 SE, 6 idiopathic
erythrocytosis) and 10 healthy controls [52]. PB and BM
EEC were positive for 81% and 84% of PV patients, and
94% of PV were diagnosed when BM and PB EEC assays
were performed with a specificity of near 100%. The diagnostic impact of low serum EPO levels (ELISA assay) was evaluated in a multicenter study including 186 patients (116 PV
according to the PVSG, 66 SE and 4 idiopathic erythrocytosis)
[60]. The majority of PV patients displayed a serum level
below the normal range of 3.3 IU/l with a sensitivity of 87%
(101/116), a specificity of 97% and a positive predictive value
of 97.8%. Statistical analysis (ROC curves) could define two
thresholds allowing a specific and correct diagnosis of PV by
EPO values below 1.4 IU/l and SE by EPO values above 13.7
IU/l. Only 66% (65/99) of PV and 20% (13/66) of SE were
diagnosed with these cut-off points, indicating an overlap of
serum EPO in PV versus control and controls versus SE [60].
Histopathology from bone marrow biopsies clearly distinguishes PV from congenital polycythemias (CP) or secondary
erythrocytosis (SE) with a sensitivity and specificity of more
than 95% to almost 100% [4044].

Pathological (P)criteria diagnostic for PV

P 1. Bone marrow pathology: increased cellularity with trilineage myeloproliferation (i.e. panmyelosis).
Proliferation and clustering of small to giant (pleiomorphic) megakaryocytes.
Absence of stainable iron.
No pronounced inflammatory reaction (plasmacytosis, cellular debris).
P 2. Bone marrow biology: spontaneous erythroid colony (EEC) formation.
P 3. Molecular biology: heterozygous or homozygous JAK2 V617F mutation.
P1 + P2 + P3 = true PV [1,2,91].
P1 or P2 plus P3 plus C1 is classical PV
P1 or P2 plus P3 plus any of C2 to C10 is early PV mimicking ET
Diagnosis PV:
C1 plus P1 and P2 or C1 and P2 plus any other criterion establish classical PV
P1 and C2 plus any other C criterion establish masked, or early PV
P1, 2, 3 plus C1, 3 plus none of the others is consistent with erythrocythemic
stage of PV

6. Myelofibrosis (MF) versus chronic idiopathic

myelofibrosis (CIMF)
Prefibrotic CIMF according to WHO [24] and ECP [25] is a
dual mixed proliferation of increased granulopoiesis and megakaryopoiesis dominated by immature giant megakaryocytes
which are conspicuously enlarged due to increase of nuclear
as well as cellular size with bulky and irregular roundishshaped nuclei, so-called cloud-like nuclei, which are never
seen in ET and PV (Table 2) [2236]. The PVSG criteria for
the diagnosis of ET include prefibrotic or early fibrotic CIMF,
which is usually associated with pronounced thrombocythemia
(false ET) (Table 4) [33,77]. Classical CIMF is defined as a
clinicopathological entity not preceded by any other MPD,
CML or MDS and characterized by various degrees of anemia,
splenomegaly, a leuko-erythroblastic blood picture with tear
drop-shaped erythrocytes and dry tap on bone marrow aspiration due to various degrees of bone marrow collagen fibrosis or
osteosclerosis (Table 8) [7076]. All studies and reports in the
literature on CIMF up to date have disregarded and excluded
just by definition the prefibrotic and early reticulin fibrotic
stage of CIMF (Table 2).
MF is not a disease because reticulin and collagen fibrosis
are produced by polyclonal fibroblasts as the consequence of
cytokines released from the clonal granulocytic and megakaryocytic proliferative cells in PV and CIMF. The well-known
Baumeister scoring system was developed on aspirated bone
marrow samples [78]. The Manoharan system scores the
degree of reticulin derived bone marrow biopsy [79]. A scoring
system based on morphometric analysis (point intersection
with an ocular grid) and quality of fibers (reticulin and collagen fibers) and the bone marrow fiber density (fine or course
reticulin and some or course bundles of collagen) have been
proposed by Georgii et al. [1113] and by Thiele et al. [18,
23]. All these different scoring systems for MF use different
criteria for grading of reticulin and collagen, are subjective
and not comparable by lack of strict criteria. A panel of experienced European pathologists and a foreign expert reached a

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J.J. Michiels et al. / Pathologie Biologie 55 (2007) 92104

consensus on how to grade bone marrow fibrosis (myelofibrosis: MF) in bone marrow biopsies of patients with CIMF or PV
[80]. Grading of MF was simplified by using four easily reproducible categories including differentiation between reticulin
and collagen [80]. According to defined standardized semiquantitative grading of reticulin and collagen fibrosis in the
bone marrow, MF can reliably be graded at the pathological
bone marrow level as 0 in prefibrotic, as 1 in early fibrotic,
as 2 in classical fibrotic and as 3 in classical sclerotic CIMF
(Tables 2 and 8) [80].
MF is not a feature of true ET and very few ET patients will
develop myelofibrosis during long-term follow-up [11,12,81
83]. MF is present in only a minority of PV patients at time of
diagnosis, but all stages of myelofibrosis have been observed
during long-term follow-up [11,12,81,82]. As compared to a
normal or near normal life expectancy in true ET and PV, prefibrotic and early fibrotic CIMF with maturation defects of
enlarged dense clustered megakaryocytes (pronounced dysmegakaryopoiesis) are featured by slowly progressive myelofibrosis (MF) progressive splenomegaly, thrombocytopenia and/or
anemia and a significantly shortened life expectancy as compared to a normal or near normal life expectancy in ET and PV
[8487]. It is reasonable to assume that the prognosis of CIMF
may depend on the grade of MF in the bone marrow at time of
diagnosis or evaluation. Early CIMF with MF grade 0 (prefibrotic) and grade 1 (early fibrotic) show a more favorable prognosis than advanced stage CIMF with grade 3 MF. However,
the survival curves of CIMF patients with grade 0, 1 and 2 are
not significantly different. Age, anemia, leukocyte and platelet
counts, but not the degree of MF (except MF grade 3 and a
hypocellular bone marrow) appeared to be the most reliable
and important parameters for prognosis and survival [8789].
The Sheffield scoring system is based on age, hemoglobin
(10g/dl) and the presence or absence of cytogenetic abnormal-

ities [89]. The Lille scoring system is derived from patients

with classical CIMF or agnogenic myeloid metaplasia
(AMM) and based on two adverse prognostic factors namely
hemoglobin < 10 g/dl and leukocytes count < 4 or
> 30 109/l88. The Lille score is widely used and able to separate patients with advanced (classical) CIMF into three groups
with low (score 0), intermediate (score 1) and high risk (score
2) of progressive disease and loss of life expectancy. The
Cologne score is derived from patients with prefibrotic, early
fibrotic and classical CIMF and uses age (70 years), hemoglobin (10 g/dl), platelets (< 300 109/l) and leukocytes
(> 20 109/l) myeloblasts (> 2%) or erythroblasts (> 2%) as
factors with prognostic impact. The Cologne scoring system
reveals a strong discriminating power and is applicable for prefibrotic and early CIMF (CIMF 0-1) as well as advanced stages
of CIMF (CIMF 2-3). Applying the Lille score to a cohort of
458 patients with prefibrotic or early fibrotic CIMF (CIMF 01), 398 belonged to the low risk group with a very good prognosis (60% survival after 15 years) and only 57 and 4 belonged
to the intermediate and high risk group with a much less favorable prognosis (less than 50% survival after 5 years) [87].
There may be an observer disagreement with regard to the
classification and natural history of prefibrotic and early CIMF
and its differentiation from true ET [87,90]. Those cases with
prefibrotic CIMF and slight dysmegakaryocytopoiesis are featured by slowly progressive myelofibrosis and splenomegaly
and have a life expectancy close to normal similar as in PV
[87]. Discussions between clinicians and pathologists reveal
that diagnostic differentiation between true ET and thrombocythemia as the presenting feature of prefibrotic MM with
slight maturation defect of enlarged clustered megakaryocytes
(slight dysmegakryopoiesis) and no or slight increased cellularity is subjective and due to a rather high inter-observer disagreement between pathologists [87,90].

Table 8
The WHO [24] and updated ECP [25,26] Criteria for the clinical diagnosis of classical fibrotic or sclerotic chronic idiopathic myelofibrosis (CIMF)
Clinical
No preceding or allied other subtype of myeloproliferative disorders ET, PV,
CML, CMML atypical CML or MDS.
Intermediate clinical stage
Anemia grade II:
hemoglobin > 10g/dl
Definitive leuko-erythroblastic blood picture and/or tear drop erythrocytes
Various degrees of thrombocythemia or normal platelet counts
Various degrees of splenomegaly
No adverse signsa
Advanced clinical stage
Anemia grade III:
hemoglobin < 10 g/l
Various degrees of thrombocytopenia
plus one or more adverse signsa
Molecular biology
JAK2 V617 positive: post-PV MM?
JAK2 V617F negative: classical CIMF?
Screen for cytogenetic abnormalities
a

Pathological
Dry tap on bone marrow aspiration is consistent with MF grade 2 or 3.
Bone marrow pathology: megakaryocytic and granulocytic myeloproliferation
and relative reduction of erythroid precursors.
Abnormal clustering and increase in atypical giant to medium sized megakaryocytes containing clumsy (cloud-like) lobulated nuclei and definitive maturation
defects.
European consensus on grading of myelofibrosis (MF) [80]
MF 2, manifest CIMF: Diffuse increase in reticulin with extensive intersections and only focal bundles of collagen. Hypercellular bone marrow
MF 3, overt CIMF: Diffuse and dense increased reticulin with extensive interactions with course bundles of collagen and significant osteosclerosis.
Hypercellular bone marrow
Osteomyelosclerosis: Slerosis, endophytic bone formation and decreased cellularity.
Endstage hypocellular bone marrow

Adverse signs: age > 70 years, hemoglobin < 10 g/dl, myeloblasts PB > 2%, erythro-normoblasts PB > 2%, leukocytosis > 2 0 109/l, thrombocytopenia < 30
0 109/l, severe constitutional symptoms, massive splenomegaly, cytogenetic abnormalities.

J.J. Michiels et al. / Pathologie Biologie 55 (2007) 92104

7. The molecular etiology of the MPDs ET PV and MF

As to the etiology of trilinear myeloproliferation in PV,
Dameshek proposed in 1950 two highly speculative possibilities: the presence of excessive bone marrow stimulation by an
unknown factor or factors, and a lack or a diminution in the
normal inhibitory factor or factors [1]. The JAK2 V617F mutation reflects a gain of function mutation, which is in line with
the concept of Dameshek [1] that all stops to blood production in the bone marrow seem to have been pulled out, which
now appears to due to one factor, the JAK2 V617F mutation
discovered Vainchenker et al. [91] and rapidly confirmed by
several investigators [9296]. The JAK2 V617F mutation
causes hypersensitivity of hematopoietic progenitor cells to
growth factors, which readily can explain the trilinear myeloproliferation and the interrelationships between ET, PV and
CIMF (Table 9) [2,9799]. Applying the PVSG criteria, 42
50% of the ET patients, 4267% of CIMF patients, 7292%
of PV patients have the mutated the JAK2 allele as detected
by DNA sequencing [9296]. A much higher frequency of
JAK2 of 97% in PV, 4957% in ET and 57% in CIMF was
described in three studies that used allele-specific polymerase
chain reaction (PCR) analysis [94,96,100,101]. The presence
of the JAK2 V617F mutation was strongly correlated with
PRV-1 over expression and the ability to form spontaneous
EEC in all three subtypes of MPD (Tables 6 and 7) [102,
103]. The JAK2 V617F mutation was absent in more than
600 healthy controls, in patients with Ph1+ CML, in patients
with reactive thrombocytosis [9296], and in patients with de
novo AML, CLL, B-ALL and T-ALL [96,104]. The mutation
has been found rarely in MDS (3/116 = 2.6%) [92,96], hypereosinophilic syndrome (2/145 = 1.4%) [92,96,105,106], but
somewhat more frequent in chronic myelomonocytic leukemia,
and atypical CML 31/408 = 7.6%) [92,96,105,106]. The JAK2
V617F mutation is rather frequent in unclassified MPD
(13/53 = 25%) [96] and frequent in AML with preceding MPD
(12/22 = 55%) [106]. The latter finding arises a key question

101

whether the JAK2 V617F mutation, homozygous in particular

or additional genetic events and/or the use of potential leukemogenic drugs contribute to the leukemic transformation of
MPDs.
The current concept is that heterozygous JAK2 V617F
mutation with increased kinase activity is enough for megakaryocyte proliferation and increased hypersensitive platelets
complicated by platelet-mediated microvascular events with
no or slightly increased erythropoiesis in ET and in early PV
mimicking ET (Table 9) [2]. Homozygous JAK2 mutation with
pronounced kinase activity is associated with trilinear megakaryocyte, erythroid and granulocytic myeloproliferation, myeloid metaplasia and secondary myelofibrosis (MF) with the
most frequent clinical picture of classical PV complicated by
major thrombosis on top of the platelet-mediated microvascular
thrombotic syndrome of thrombocythemia (Table 9). The
sequential occurrence of heterozygous and homozygous
V617F mutation can readily explain the progression of ET
and early PV to overt PV and may explain at least in part the
accompanied granulocytic proliferation, leukocyte activation
(increased LAP score and PRV-1 expression), leukocytosis
(leukocythemia), and secondary myelofibrosis (Table 9) [1,2].
Depending on the set of laboratory tests and diagnostic criteria used, the population of the MPD patients defined as ET,
PV and CIMF are heterogeneous at the molecular and biological level (Tables 1, 2, 7, 8). As the JAK2 V617F mutation is
the cause of a distinct trilinear MPD in its manifold clinical
manifestations during long-term follow-up [2], the specificity
of a positive JAK2 V617F PCR test for the diagnosis of
MPD is high (near 100%), but only half of ET and CIMF (sensitivity 50%) and the majority of PV (sensitivity 8597%) are
JAK2 V617F positive. Kiladjian et al. [107] searched for the
JAK2 V617F mutation in a cohort of 44 female ET patients
with previously documented clonality data (X-chromosome
inactivation pattern: XCIP assay). The mutation was found in
four of the nine patients with polyclonal hematopoiesis (probably due to better sensitivity of mutation detection than XCIP
assays), in 13 of the 31 patients with monoclonal hematopoi-

Table 9
Molecular etiology of platelet-mediated microvascular thrombosis, increased red cell mass and secondary myelofibrosis in JAK2 V617F positive MPDs ET, PV, MF
Vainchenker & Michiels 2005
JAK2 V617F gain of function mutation in trilinear hematopoietic cells of MPD patients is detectable in platelets, erythroblast and granulocytes
Step 1 V617F+
Step 12 V617F++
Step 12 V617F++
LOH
LOH

Spontanuous

Spontanuous

Myeloid Metaplasia: MM
CFU-MK / EEC
EEC, CFU-MK
Leukocyte activation
ET
PV
PVR-1 = LAF

Increase of enlarged
Increase of hematocrit to above 0.45-0.50: PV
Leukocythemia/cytokines
hypersensitive platelets
higher platelets
Fatigue, splenomegaly, unclassified
Already at platelet >400
MMM

Clinical Step 1

Clinical Step 2

Clinical Step 3
Microvascular

Macrovascular

Secondary MF: 30%

Thrombosis
Thrombosis
Thrombosis Thrombosis
Constitutional symptoms
Aspirin sensitive
Aspirin/Phlebotomy
IFN/Hydroxyurea
See figures 1 and 2 regarding the natural history of true PV according to Wasserman and Thiele.

MPD,

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J.J. Michiels et al. / Pathologie Biologie 55 (2007) 92104

esis and in two of the four patients with undetermined clonality

status due to non-random XCIP. These results show that the
majority of patients with ET according to the PVSG without
V617F JAK2 mutation have nevertheless a clonal hematopoietic stem cell disease. Combining results of JAK2 sequencing
and XCIP assays allowed demonstration of a clonal proliferation in 90% of ET cases. Buck et al. [108] observed a lower
incidence of 25% JAK2 V617F positivity in patients with true
ET according to the WHO and ECP criteria (personal communication). In a large prospective study of 806 ET patients diagnosed according to the PVSG criteria, about half of the ET
patients (53%) were JAK2 V617F positive. ET patients positive for the JAK2 V617F mutation have higher hemoglobin
level and a significantly higher rate of transformation into PV
during follow-up as compared to ET patients negative for the
JAK2 V617F mutation [108]. Comparing the laboratory features of JAK2 V617 positive ET and JAK2 V617 negative
ET patients in the PT-1 study clearly showed that JAK2
V617 positive ET is featured by higher values for hemoglobin,
hematocrit, neutrophil counts, lower values for serum EPO
levels, serum ferritine and MCV, and hypercellularity of the
bone marrow in biopsy material indicating that JAK2 V617
positive ET patients diagnosed according to the PVSG criteria
represent a forme fruste of PV [108] consistent with early PV
mimicking ET (ECP stage 1 in Table 6). In contrast, the JAK2
negative ET patients had significantly higher platelet counts
and showed a clinical picture of true ET with normal serum
EPO levels and increased of clustered megakaryocytes in a
normocellular bone marrow consistent with a diagnosis of
true ET (Table 1). These clinical data indicate that JAK2
V617F mutation defines one disease with several sequential
steps of ET, PV and MF during long-term follow-up (Table 9).
Patients with either JAK2 V617F positive or negative prefibrotic and fibrotic CIMF may represent two distinct entities with a
related etiology for one and the same disorder. Bone marrow
histopathology when used in combination with specific markers like JAK2 V617F, serum EPO, PRV-1, EEC, peripheral
blood parameters has a high sensitivity and specificity (almost
100%) to detect each of the early stages of MPD and to differentiate between ET, PV and CIMF in both JAK2 V617F positive and negative MPDs.

[7]
[8]

[9]

[10]

[11]

[12]
[13]

[14]

[15]

[16]

[17]

[18]

[19]
[20]

[21]

[22]

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