Myelofibrosis (MF) generally refers to a myeloproliferative neoplasm that is induced by mutations affecting the maturation, differentiation, and function of hematopoietic stem cells. These mutations may be inherited, giving rise to familial myelofibrosis, or acquired, as is the case with other, more common variants of primary myelofibrosis. MF may also be the natural progression of a different myeloproliferative disease, like polycythemia vera or essential thrombocytosis. The respective patients are diagnosed with secondary myelofibrosis. Finally, reactive myelofibrosis may occasionally be observed in those with non-myeloproliferative cancers and many other conditions, such as endocrine disorders, autoimmune diseases, and infections.
Presentation
MF presentation is diverse. About one-third of MF patients remain asymptomatic for prolonged periods of time, while others develop debilitating constitutional symptoms and signs of bone marrow failure [1]. Regarding the former, fatigue, fever, dyspnea, night sweats, and weight loss are most commonly reported. Patients may become cachectic. Additional symptoms are usually related to hematological abnormalities: There is a proliferation of predominantly megakaryocytes and granulocytes in the bone marrow, which may give rise to leukocytosis and thrombocytosis, along with circulatory disorders due to thrombophilia, thrombosis, and disseminated intravascular coagulation [2].
As the disease progresses, the reactive deposition of fibrous connective tissue increasingly interferes with hematopoiesis, and patients develop cytopenias. Anemia is common in MF patients, may increase fatigue and weakness, induce pallor and cyanosis. Anemic patients may also claim headaches and dizziness. Leukopenia, on the other hand, predisposes to infections. Thrombocytopenia is less frequently observed than thrombocytosis and renders patients prone to bleeding.
In an intent to compensate for bone marrow failure, extramedullary hematopoiesis is stimulated, entailing hepatomegaly, splenomegaly, and possibly bone pain and osteosclerosis. Besides abnormal laboratory results, palpable organomegaly may indeed be the only finding on clinical examination.
Immune System
- Splenomegaly
Splenomegaly is one of the major clinical manifestations of MF and is directly linked to splenic extramedullary hematopoiesis (EMH). [ncbi.nlm.nih.gov]
Similarly, their use in patients with splenomegaly due to splanchnic vein thrombosis in which the mechanism of splenomegaly is probably different from that of typical myelofibrosis is warranted as well. [jhoonline.biomedcentral.com]
Splenomegaly [ 1 ] First-line: hydroxycarbamide (in the absence of cytopenias). [patient.info]
- Massive Splenomegaly
However, splenectomy can be useful in selected situations—e.g., in patients with massive splenomegaly and refractory cytopenias not responsive to standard therapies. [journals.lww.com]
"With massive splenomegaly, patients develop abdominal discomfort, early satiety and weight loss," said Gotlib. In the study, many patients who received the drug experienced a significant reduction in spleen volume and a lessening of symptoms. [medicalxpress.com]
“With massive splenomegaly, patients develop abdominal discomfort, early satiety and weight loss,” said Gotlib. In the study, many patients who received the drug experienced a significant reduction in spleen volume and a lessening of symptoms. [med.stanford.edu]
[citation needed] Splenectomy is sometimes considered as a treatment option for patients with myelofibrosis in whom massive splenomegaly is contributing to anaemia because of hypersplenism, particularly if they have a heavy requirement for blood transfusions [en.wikipedia.org]
Entire Body System
- Constitutional Symptom
Ruxolitinib is highly effective in ameliorating constitutional symptoms and reducing spleen size in JAK2 -mutated and—wild type patients alike. [journals.lww.com]
The JAK inhibitors are effective in both JAK2-positive and JAK2-negative MF; one of them, ruxolitinib, is the current best available therapy for MF splenomegaly and constitutional symptoms. [ncbi.nlm.nih.gov]
Management of constitutional symptoms in PMF is very difficult and there is no evidence of benefit for any of the conventional treatments for PMF. [patient.info]
- Pathologist
Even grading the degree of fibrosis is something that is not tremendously well standardized, and there can be disagreements between pathologists. [targetedonc.com]
Introduction Since 1976, a group of European pathologists described a subtype of chronic myeloproliferative neoplasms (MPN) that was characterized by megakaryocytic and granulocytic proliferation, with atypical megakaryocyte morphology, but without any [nature.com]
All patients had BM biopsy at diagnosis or had BM samples taken at diagnosis reviewed by our pathologist (UM). On this sample, the degree of BM cellularity in paraffin-embedded BM biopsy specimens stained with hematoxylin and eosin was estimated. [journals.plos.org]
A pathologist views the bone marrow, blood, and bone under a microscope to look for abnormal cells. Enlarge Bone marrow aspiration and biopsy. After a small area of skin is numbed, a bone marrow needle is inserted into the patient’s hip bone. [cancer.gov]
Respiratoric
- Cough
[…] full before finishing a meal (because of an enlarged spleen) Bone pain Easy bleeding, bruising Fatigue Increased likelihood of getting an infection Pale skin Shortness of breath with exercise Weight loss Night sweats Low grade fever Enlarged liver Dry cough [nlm.nih.gov]
Over time you might notice: Major weight loss Itching Coughing up blood Seizures Pressure on your spinal cord Bone and joint pain Easy bleeding that could make surgery risky Pain and swelling in your joints, called gout Some people can also get acute [webmd.com]
These tumors may cause problems such as bleeding in your gastrointestinal system, coughing or spitting up of blood, compression of your spinal cord, or seizures. Bleeding complications. [mayoclinic.org]
- Dry Cough
cough Itchy skin The health care provider will perform a physical exam and ask about the symptoms. [nlm.nih.gov]
cough Itchy skin Exams and Tests The health care provider will perform a physical exam and ask about the symptoms. [m.ufhealth.org]
Gastrointestinal
- Abdominal Pain
Year-Old Man Diagnosed With Primary Myelofibrosis December 2018 A 72-year old man presents to primary care physician with complaints of fatigue, headache, night sweats, poor appetite, and 10-15lb weight loss over past few months; report of increased abdominal [targetedonc.com]
The most common grade 3 or worse adverse events were anaemia (14 [14%] of 104 in the momelotinib group vs seven [14%] of 52 in the BAT group), thrombocytopenia (seven [7%] vs three [6%]), and abdominal pain (one [1%] vs three [6%]). [ncbi.nlm.nih.gov]
MF patients can also suffer from debilitating constitutional symptoms, such as drenching night sweats, fatigue, severe itching (pruritus), abdominal pain, fever and bone pain. [geron.com]
In turn, this causes the liver and spleen to become enlarged and creates some of the symptoms such as abdominal pain, feeling full, weight loss, fatigue, itching, bone pain, and night sweats that people diagnosed with MF may experience. [cancercare.org]
- Early Satiety
The WG focused on the seven core symptoms of MF: fatigue, night sweats, pruritus, abdominal discomfort, pain under the ribs on the left side, early satiety, and bone pain. [ncbi.nlm.nih.gov]
The spleen continues to enlarge, which leads to early satiety. Painful episodes of splenic infarction may occur. The patient becomes cachectic and may experience severe bone pain, especially in the upper legs. [accessmedicine.mhmedical.com]
It could be the presentation of symptoms: Unexpected weight loss, sometimes early satiety. “I’m filling up early when I eat.” It could be a pronounced fatigue or night sweats. [targetedonc.com]
satiety, and abdominal discomfort. [journals.lww.com]
- Abdominal Fullness
Symptoms may include any of the following: Abdominal fullness, pain, or feeling full before finishing a meal (because of an enlarged spleen) Bone pain Easy bleeding, bruising Fatigue Increased likelihood of getting an infection Pale skin Shortness of [nlm.nih.gov]
Patients most commonly present with fatigue due to anemia or abdominal fullness related to splenomegaly. Uncommon presentations include bleeding and bone pain. On examination, splenomegaly is almost invariably present and is commonly massive. [accessmedicine.mhmedical.com]
A sense of abdominal fullness caused by enlargement of the spleen Bone pain Bruising and easy bleeding due to declining platelets Fatigue Increased susceptibility to infection Pallor and shortness of breath related to anemia Hyperuricemia (a high level [cancer.columbia.edu]
Symptoms Symptoms may include any of the following: Abdominal fullness, pain, or feeling full before finishing a meal (because of an enlarged spleen) Bone pain Easy bleeding, bruising Fatigue Increased likelihood of getting an infection Pale skin Shortness [m.ufhealth.org]
- Diarrhea
CASE PRESENTATION: We report a 36-year-old pregnant woman who presented with cytopenia, fatigue, vomiting, and diarrhea for 20 days on the background of newly diagnosed myelofibrosis secondary to gastric signet ring adenocarcinoma. [ncbi.nlm.nih.gov]
Besides abdominal swelling, other symptoms such as digestive disorders (diarrhea and constipation) heartburn, and indigestion triggered by fatty foods may also appear. [aoporphan.com]
The most common side effects of Jakafi include: for certain types of MF and PV – low platelet or red blood cell counts, bruising, dizziness, headache, and diarrhea; and for acute GVHD – low platelet, red or white blood cell counts, infections, and fluid [jakafi.com]
Jakafi may cause nausea, diarrhea, liver inflammation, reduced platelets, and increased cholesterol in some. [verywellhealth.com]
Skin
- Night Sweats
So, malfunctioning bone marrow, anemia—low platelets in some patients—and low white cell count coupled with a very large spleen, enlargement of the liver, and very poor quality of life—bone aches and pains, night sweating, low-grade fevers, itching, night [onclive.com]
Other symptoms may include: Fatigue A general feeling of discomfort Difficulty breathing Weight loss Fever and night sweats Anemia Abnormal bleeding Patients with myelofibrosis have an increased risk of bleeding. [stanfordhealthcare.org]
Case: 72-Year-Old Man Diagnosed With Primary Myelofibrosis December 2018 A 72-year old man presents to primary care physician with complaints of fatigue, headache, night sweats, poor appetite, and 10-15lb weight loss over past few months; report of increased [targetedonc.com]
[…] fatigue, fever, night sweats and weight loss. [mpnresearchfoundation.org]
- Erythema
Thalidomide is US Food and Drug Administration (FDA)-approved for use in acute erythema nodosum leprosum and, in combination with dexamethasone, in newly diagnosed myeloma. [doi.org]
[…] per week in patients with high-risk β-thalassemia, it can reverse iron-induced cardiac dysfunction and increase long-term survival. 90 However, treatment-related side effects include infusion site discomfort (nearly 100%, with the development of local erythema [journals.sagepub.com]
Workup
The World Health Organization has defined diagnostic criteria for primary MF and distinct myeloproliferative neoplasms that may ensue fibrotic changes of the bone marrow [3]. Due to the diversity of conditions that may be associated with reactive myelofibrosis, there are no common guidelines on this matter. In general, the diagnosis of MF is based on morphologic features of the bone marrow, molecular and genetic findings, along with laboratory data:
- The examination of bone marrow biopsy specimens should reveal megakaryocytic proliferation and megakaryocytic atypia, which are usually accompanied by reticulin or collagen fibrosis. In the prefibrotic phase of primary MF, reticulin and collagen fibrosis may be absent, but increased bone marrow cellularity with granulocytic proliferation and reduced erythropoiesis may be observed [4].
- Genetic studies should be realized to determine the presence of driver mutations of genes JAK2, CALR, and MPL. Due to the high prevalence of the JAK2 V617F mutation, first analyses generally aim at the detection of this gene defect. In the absence of driver mutations, samples may be tested for associated somatic mutations affecting genes ASXL1, EZH2, TET2, IDH1/2, SRSF2, or SF3B1, among others [1] [5].
- In order to distinguish primary MF from secondary MF, criteria established for the diagnosis of polycythemia vera, essential thrombocytosis, other myeloid neoplasms, and myelodysplastic syndromes should be reviewed. Primary MF is only to be diagnosed if these criteria are not met. Additionally, reactive myelofibrosis developing in the setting of lymphoid or metastatic malignancy, infection, inflammation, or toxic myelopathy should be ruled out [3].
- Minor criteria for the diagnosis of MF comprise anemia, leukocytosis, increased serum lactate dehydrogenase, leukoerythroblastosis (with teardrop-shaped red blood cells), and palpable splenomegaly.
Ultrasound
- Enlargement of the Liver
My husband was diagnosed with MF in 2004, no symptoms, JAK neg, just enlarged spleen and liver. Everything was relatively fine, but we were upset that there was no help until symptoms start to show. That's where we are now. [csn.cancer.org]
The resulting fibrous scar tissue formation leads to severe anemia, weakness, fatigue and an enlarged spleen and liver. Is a type of chronic leukemia and can occur on its own (primary myelofibrosis) or as a result of another bone marrow disorder. [llscanada.org]
The disease can lead to an enlarged spleen and liver. The origins and risk factors for myelofibrosis are unknown. The condition is rare, affecting 3,000 to 4,000 people in the U.S. each year. [cancercare.org]
[…] and liver bone pain gout loss of appetite and weight loss fever and night sweats Tests to diagnose myelofibrosis You have a blood test to check your blood cell count. [cancerresearchuk.org]
The result is extensive scarring in your bone marrow, leading to severe anemia, weakness, fatigue, and often, an enlarged spleen and liver.'' [cmleukemia.com]
Serum
- Thrombocytosis
BM Gomori silver impregnation (400×) Primary myelofibrosis, overt fibrosis. 72-year old man, leucocytosis 16×109/L, thrombocytosis 767×109/L, JAK2 unmutated. Intense reticulin fibrosis (MF-2 according to the EUMNET classification). [leukemia-cell.org]
Myelofibrosis can also occur as a secondary process following the other myeloproliferative disorders (eg, polycythemia vera, essential thrombocytosis). [accessmedicine.mhmedical.com]
Thrombocytopenia is less frequently observed than thrombocytosis and renders patients prone to bleeding. [symptoma.com]
Hydroxyurea has traditionally been the preferred and most commonly used agent - moderately effective at improving splenomegaly, leukocytosis and thrombocytosis [ 5 ]. [patient.info]
- Erythrocytosis
We describe how treatment with a JAK1/2 inhibitor, ruxolitinib, led to the rapid amelioration of marrow fibrosis, erythrocytosis and thrombocytopenia in a 77-year-old man with post-PV MF who carried a JAK2 exon 12 mutation (JAK2H538QK539L). [ncbi.nlm.nih.gov]
[…] virus-like particles, granulocytic and erythrocytic hyperplasia, chromosomal abnormalities–eg, of 21q which may correlate with reverse transcriptase activity Lab Thrombocytosis–1-14 x 1012/L with giant and bizarre forms, hypochromic and microcytic anemia or erythrocytosis [medical-dictionary.thefreedictionary.com]
JAK2 exon 12 mutations in polycythemia vera and idiopathic erythrocytosis. N Engl J Med 2007; 356 : 459–468. 32. Pikman Y, Lee BH, Mercher T, McDowell E, Ebert BL, Gozo M et al. [doi.org]
Treatment
It is of particular importance to determine the etiology of MF and to adapt the therapeutic regimen to the individual case. Reactive MF may be induced by treatable diseases and tend to have a much better outcome than cases related to myeloproliferative neoplasms [6]. Therapeutic options are limited for patients with primary MF or MF related to other myeloproliferative neoplasms.
JAK 1/2 inhibitor ruxolitinib has received approval for MF treatment in 2012 and continues to be the only drug licensed to this end [7]. Ruxolitinib offers a significant benefit in relieving symptoms and reducing splenomegaly, but there is an inherent risk of worsening cytopenias that requires close monitoring of hematological parameters and possibly pharmacological countermeasures [6]. Danazol, erythroid-stimulating agents, and immunomodulatory drugs may be administered to control ruxolitinib-induced anemia, but the results remain unsatisfactory [7].
Several other JAK inhibitors have been or are currently examined in clinical studies, e.g., fedratinib, pacritinib, momelotinib, and itacitinib. While some have been withdrawn from development due to severe side effects, others have proven to be well tolerated, highly effective, and less myelosuppressive than ruxolitinib [1]. New concepts of MF treatment may be of particular interest to those when ruxolitinib has to be discontinued, as this decision is generally linked to a dismal outcome [7].
To date, hematopoietic stem cell transplantation remains the only curative option. There are comprehensive guidelines concerning patient eligibility, the use of JAK inhibitors before and after the transplantation, conditioning regimens, and donor selection. In general, patients with increased or high-risk disease and age <70 years should be considered potential candidates for stem cell transplantation, while those with intermediate-risk MF and age <65 years should be proposed as candidates only if they present with either refractory, transfusion-dependent anemia, or a percentage of blasts in peripheral blood >2%, or adverse cytogenetics [5]. In those with low-risk disease, transplant-related morbidity and mortality outweigh the possible benefits of stem cell transplantation [1]. The respective definitions are part of the International Prognostic Scoring System and its extensions and can be looked up in the literature.
Prognosis
The Dynamic International Prognostic Scoring System is widely used to define the prognosis of an individual patient and considers a total of eight predictors: age >65 years, hemoglobin level <10 g/dl, leukocytes >25*10^9/l, platelet count <100*10^9/l, circulating blasts >1%, constitutional symptoms, transfusion dependency, and unfavorable karyotype [8]. An even better prognostication may be achieved by additionally considering morphological data, most importantly the grade of bone marrow fibrosis [3]. Median survival times of primary MF patients are estimated at 6 years but may range from months to several years, which emphasizes the need for individual assessments [8].
Besides progressive marrow failure and complications from thrombosis, hemorrhages, infections, and portal hypertension, leukemic transformation has been identified as a common cause of death and has, in turn, been related to severe thrombocytopenia and unfavorable karyotypes. In this context, an unfavorable karyotype or platelet count <100*10^9/l may imply a 10-year risk of leukemic transformation of up to 31% [8].
Etiology
According to the current classification of the World Health Organization, primary MF is a clonal myeloproliferative neoplasm, as are Philadelphia chromosome-positive chronic myeloid leukemia, chronic neutrophilic leukemia, chronic eosinophilic leukemia, polycythemia vera, essential thrombocythemia, and mastocytosis [4]. Secondary MF may develop as a complication of any of these diseases, but is most commonly seen in patients diagnosed with polycythemia vera and essential thrombocythemia.
All these conditions - MF as well as other myeloproliferative neoplasms - have been related to mutations of the JAK2, CALR, and MPL genes. JAK2 mutations are the most common driver mutations in MF patients and are identified in little more than half of all cases, while abnormalities of CALR are determined in less than one-third of the affected individuals. MPL mutations account for less than 10% of MF cases, and a similar share of cases is classified as triple-negative. Other, as-of-yet unknown driver mutations are assumed to trigger MF in these patients [1]. It shall be stressed again that none of these mutations is specific for MF. Just to take the example of JAK2 V617F, which is determined in nearly half of MF cases: This same mutation accounts for a similar percentage of essential thrombocythemia and as much as 90% of cases of polycythemia vera [4].
MF driver mutations eventually alter cytokine release by hematopoietic cells and their precursors. Notwithstanding, cytokines promoting bone marrow fibrosis may originate from non-myeloid cells or iatrogenic sources, which must then be considered as triggers of reactive MF. Kuter and colleagues have compiled an extensive, yet not complete list of conditions associated with increased bone marrow fibrosis [9].
Epidemiology
The annual incidence of MF has been estimated at 0.1-1 per 100,000 individuals. Most patients present during their sixth or seventh decade of life, and only about 10% of all cases are diagnosed in individuals aged <40 years [1] [3]. MF in children is increasingly rare but may occur in the setting of germline mutations and familial MF [10]. Regardless of age, males and females are affected equally.
Pathophysiology
MF is caused by the proliferation of clonal hematopoietic stem cells. Their proliferative advantage over normal cells is conferred by distinct mutations entailing the constitutive activation of the JAK-STAT pathway. This applies to all patients, regardless of whether they carry any of the known driver mutations or are classified as triple-negative [1]. Increased signaling via the JAK-STAT pathway interferes with the regulation of cytokine release and thus contributes to the creation of a proinflammatory and fibrogenic milieu. In detail, vascular endothelial growth factor, transforming growth factor-β, interferon-γ, interleukins 2, 8, and 17, and lipocalin-2 are believed to act in concert to promote the deposition of fibrous connective tissue. At the same time, the population of clonal cells continues to grow, occupying more and more space. Both mechanisms - bone marrow fibrosis and overgrowth of clonal cells - progressively impede polyclonal hematopoiesis and ultimately lead to a state of bone marrow failure and cytopenias [6].
Prevention
Owing to persisting knowledge gaps regarding the triggers of stem cell degeneration, no recommendations can be given to prevent the onset of primary MF. Concerning MF following essential thrombocythemia and polycythemia vera, novel therapeutics aimed at the prevention of disease progression to myelofibrosis are the focus of current clinical trials [11]. Things look much better with regard to reactive MF: Here, the adequate management of the underlying condition underlying may indeed delay or prevent bone marrow fibrosis.
Summary
MF is often used as a synonym of primary MF but is actually a descriptive term referring to the presence of bone marrow fibrosis and extramedullary hematopoiesis [3]. These may be observed in a wide range of diseases, including malignancies, endocrine disorders, autoimmune diseases, and infections, which may or may not originate in the bone marrow [6]. By far the most common variant of MF is primary MF, and the second most important cause of the disease are hematological malignancies. The latter may follow a more aggressive course during their later phases and transform to overt MF. Finally, there's a third group of MF cases that may be classified as reactive bone marrow fibrosis. A number of heterogeneous disorders fall into this category, all of which are uncommon etiologies of bone marrow fibrosis.
Patient Information
Myelofibrosis (MF) denotes the progressive deposition of fibrous tissue in the bone marrow. This condition may have different causes but is most commonly induced by acquired mutations of hematopoietic stem cells, the triggers of which remain unknown. The respective patients are diagnosed with primary MF and are usually of advanced age. Another share of patients suffers from secondary MF, which may be considered an advanced stage of myeloproliferative neoplasms such as essential thrombocythemia and polycythemia vera. Finally, MF may be caused by autoimmune diseases, inflammation, infection, endocrine imbalances, and many other disorders. These patients account for a very small share of MF cases, though.
Both the overgrowth of mutated cells and progressive bone marrow fibrosis interfere with the formation of new, healthy blood cells. MF patients may thus present with distinct hematological abnormalities, such as increased or decreased counts of red blood cells, white blood cells, and platelets. They may claim fatigue and weakness due to anemia, be predisposed to infections, thrombosis, or bleeding. As the body tries to compensate for bone marrow failure, hematopoiesis is stimulated elsewhere, entailing the enlargement of liver and spleen.
None of these symptoms is specific for MF, and a thorough workup is required to diagnose this disease and to identify possible causes. Both treatment and prognosis largely depend on many factors, e.g., the age of the patient, the presence of comorbidities, the stage of the disease and the precise mutations underlying MF. The median survival of patients diagnosed with primary MF, the most common form of the disease, has been estimated at 6 years but may range from months to several years.
References
- O'Sullivan JM, Harrison CN. Myelofibrosis: clinicopathologic features, prognosis, and management. Clin Adv Hematol Oncol. 2018; 16(2):121-131.
- Thiele J. Philadelphia chromosome-negative chronic myeloproliferative disease. Am J Clin Pathol. 2009; 132(2):261-280.
- Swerdlow SH, Campo E, Harris NL. WHO Classification of Tumours of Haematopoietic and Lymphoid Tissues 4ed. Lyon, France: IARC Press; 2017.
- Vardiman JW, Thiele J, Arber DA, et al. The 2008 revision of the World Health Organization (WHO) classification of myeloid neoplasms and acute leukemia: rationale and important changes. Blood. 2009; 114(5):937-951.
- McLornan DP, Yakoub-Agha I, Robin M, Chalandon Y, Harrison CN, Kroger N. State-of-the-art review: allogeneic stem cell transplantation for myelofibrosis in 2019. Haematologica. 2019; 104(4):659-668.
- Marcellino B, El Jamal SM, Mascarenhas JO. Distinguishing autoimmune myelofibrosis from primary myelofibrosis. Clin Adv Hematol Oncol. 2018; 16(9):619-626.
- Bose P, Alfayez M, Verstovsek S. New Concepts of Treatment for Patients with Myelofibrosis. Curr Treat Options Oncol. 2019; 20(1):5.
- Gangat N, Caramazza D, Vaidya R, et al. DIPSS plus: a refined Dynamic International Prognostic Scoring System for primary myelofibrosis that incorporates prognostic information from karyotype, platelet count, and transfusion status. J Clin Oncol. 2011; 29(4):392-397.
- Kuter DJ, Bain B, Mufti G, Bagg A, Hasserjian RP. Bone marrow fibrosis: pathophysiology and clinical significance of increased bone marrow stromal fibres. Br J Haematol. 2007; 139(3):351-362.
- Sieff CA, Malleson P. Familial myelofibrosis. Arch Dis Child. 1980; 55(11):888-893.
- Aruch D, Mascarenhas J. Contemporary approach to essential thrombocythemia and polycythemia vera. Curr Opin Hematol. 2016; 23(2):150-160.