الفهرس 
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Abstract
The chronic myeloproliferative diseases are a group of conditions characterized by unregulated blood cell production, that is due either to excessive numbers of erythrocytes, leukocytes or platelets or their defective function or apoptosis.
Chronic myelogenous leukemia t(9;22)(q54:94) is BCR/ABL positive polycythemia vera, essential thrombocythemia and primary myelofibrosisare negative for Philadelphia chromosome and have overlapping clinical features but exhibit different natural histories and different therapeutic requirements.
The aim of the study was to study the role of angiogenesis and its significance in myeloproliferative disorders.
The study included 30 patients with myeloproliferative neoplasms presenting to the Haematology Department, Medical Research Institute before treatment, 15 of which had CML, 11 patients had PMF, two patients with PV and 2 with ET. There were seventeen males and thirteen females aged from 22-81 years. The study also included 10 lymphoma patients with normal bone marrow as a control group.
All individuals included in the study were subjected to the following:
•Detailed history taking.
•Thorough clinical examination to assess the presence of hepatomegaly, splenomegaly, and / or lymph node enlargement.
•Radiological investigations
oPlain x ray chest.
oAbdominal ultrasound.
oComputed tomogaphy of the abdomen.
•Laboratory investigations including:
a- Complete blood picture.
b- Bone marrow aspiration and bone marrow trephine biopsy.
c- Immunohistochemical staining of bone marrow blood vessels using monoclonal antibodies to visualize anti vWF and anti- thrombomodulin as endothelial cell markers. Hot spot areas were defined as areas with the maximum number of microscopic counts using low power magnification. The average of the number of bone marrow blood vessels in four hot spot areas was taken by two investigators independently. Areas of staining with no defined lumen were counted as a single vessel.
d- Immunohistochemical study of VEGF expression in bone marrow trephine biopsy sections (VEGF intensity of staining, percentage of positive cells and IRS) were recorded. Intensity of immunocytochemical procedure was evaluated using the ImmunoReactive Score (IRS). The applied scale took into account both intensity of the color reaction and precentage of cells, which exhibited the positive reaction. Final result represented the product of the two parameters and its value ranged between 0-12.
The most common presentation was fatigue, abdominal discomfort and weight loss. Splenomegaly was present in 22 of 30 cases while hepatomegaly was found in 5 cases only
CBC results showed that the mean hemoglobin concentration of CML patients was with a mean of 10.8±1.9 g/dL while in PMF was 7.89±3.2 g/dL. In ET, hemoglobin concentration was 11.95±1.48.The hemoglobin level of for the PRV patients was 18.7 g/dL for one case and 19.0 g/dL for the other, while in ET, the hemoglobin level was 10.9 g/dL in one case and 13.0 g/dL in the other case.
As regards total leukocytic count, it was significantly increased in CML cases with a mean of 134.3±74.3x109/L. In PMF, the mean was 9.7±8.2x109/L,while in PRV patients it was 9 ×109/L in one case and 12 ×109/L in the other, and in the two ET cases, it was 10 ×109/L and13 ×109/L respectively.
As regards, the platelet count, it was significantly increased in ET patients with a mean of 870±170.0x109/L, while in CML the platelet count was 348.7±180.5x109/L. In PMF, the mean platelet count was 145.4±154.6x109/L.In PRV patients, the platelet count was 180 ×109/L in one case and 210 ×109/L in the other, and in the two ET cases, it was 750 ×109/L and 990 ×109/L respectively.
As regards bone marrow trephine biopsies of the studied groups, seventeen of the the thirty CMPDs patients showed marked bone marrow cellularity, five patients showed moderate cellularity, while eight patients showed mild cellularity. No correlation was found with VEGF expression or MVD.
As regards, bone marrow fibrosis, eight of the the thirty patients showed mild fibrosis, six patients showed moderated fibrosis, eleven patients showed marked fibrosis, while five patients with no fibrosis. The pattern of bone marrow infiltration was diffuse in 86.67% of cases while it was patchy in 13.5% of them.
There was a significant increase in the number of bone marrow blood vessels in MPNs on using anti vWF (t=8.07, p<0.01), CML (t=8.37, p<0.01), PMF (t=7.8, p<0.01) with the means of (10.9±3.3, 10.9±2.19 and 12.9±3.1/H.P.F. respectively) as compared to the control group (mean=4.8±1.48/H.P.F.).
On using anti TM antibodies as endothelial cell markers, there was also a statistically significant increase in BM blood vessels in MPNs (t=8.56, p<0.01), CML (t=8.98, p<0.01), PMF (t=9.2, p<0.01) with the means of (10.4±3.1, 10.07±1.87 an 12.5±2.6/H.P.F. respectively) as compared to the control group with a mean of 4.6±1.2/H.P.F.
There was a positive correlation between the number of bone marrow blood vessels counted using immunohistochemical staining with anti-vWF and anti-thrombomodulin antibodies as endothelial cell markers.
VEGF expression (percentage of positive cells) was statistically increased in MPNs (t=18.53, p<0.01), CML (t=15.17, p<0.01), PMF (t=13, p<0.01)(with the means of 48.6±12.2, 48.2±10.5 and 54±12 respectively) when compared to to the control group (mean=6.3±1.6) Immunoreactive score (VEGF expression) was also increased in MPNs, CML, PMF as compared to the control group.
Inflammatory mononuclear cells (fibroblasts, macrophages, polymorphs) were also stained with VEGF. These cells were found as few isolated or clustered elements throughout the stroma of the bone marrow, and were clearly distinguishable from tumor cells both on morphological basis and because they usually displayed a different staining intensity (in plus or minus) from tumor cells. Care was taken to recognize these inflammatory cells and omit them from the evaluation.
There was a positive correlation between VEGF expression and the number of bone marrow blood vessels counted by immunohistochemical staining with both anti-vWF and anti-thrombomodulin antibodies.
PMF patients showed a higher level of VEGF expression and an increased number of bone marrow blood vessels than other MPNs patients but this was not statistically significant. The present study also revealed that most cases with increased angiogenesis and VEGF expression had splenomegaly.
In conclusion, the present study has demonstrated an increase in bone marrow blood vessels visualized by immunohistochemical staining with anti-vWF and anti-thrombomodulin and the increased expression of VEGF which is a common feature of myeloproliferative neoplasms especially in patients with PMF. VEGF expression is also a more precise method than measuring VEGF in serum or plasma levels. Our data emphasize the role of increased angiogenesis. It could be of value as a prognostic marker of disease progression as well as response to therapy.