Constantina A. Pappa

Levels of serum cytokines and acute phase proteins in patients with essential and cancer-related thrombocytosis.

&NA; Essential thrombocytosis (ET) is a myeloproliferative disorder resulting in an increased production of abnormal platelets. Reactive thrombocytosis (RT) is occasionally observed in clinical situations including chronic inflammation and malignancy. The aim of the present study was to evaluate the discriminatory efficiency of various laboratory tests in patients with ET and cancer‐related RT. Forty‐five patients with ET, 52 patients with RT, and 25 age‐matched normal individuals comprised the study population. Plasma interleukin‐1 alpha (IL‐1a), IL‐2, IL‐6, tumor necrosis factor alpha (TNF‐a), platelets, hematocrit, hemoglobin, erythrocyte sedimentation rate (ESR), C‐reactive protein (CRP), lactate dehydrogenase (LDH) and ferritin were determined. We found increased levels of ferritin, LDH, CRP, ESR, IL‐1a, and IL‐6 in RT compared with ET (p < 0.01 to p < 0.0005). Hemoglobin, hematocrit, and platelets were significantly lower in RT than in ET (p < 0.0005). Furthermore, ferritin and ESR were negatively correlated with Hct, hemoglobin, and TNF‐a, whereas ferritin was positively correlated with ESR, IL‐1a, IL‐6, and CRP, and IL‐1a was positively correlated with IL‐6. We consider that the aforementioned parameters should be included in the investigation of unexplained thrombocytosis for the differentiation of essential from cancer related thrombocytosis.

The relation between bone marrow angiogenesis and the proliferation index Ki-67 in multiple myeloma

Aim: Angiogenesis correlates with disease progression in various haematological malignancies. This study investigated the association between microvascular density (MVD) and the Ki-67 proliferation index (Ki-67 PI), bone marrow infiltration, and C reactive protein (CRP) in patients with multiple myeloma. Methods: Bone marrow MVD was examined in 44 biopsies at diagnosis and 15 in plateau phase by immunostaining the endothelial cells with a monoclonal antibody to CD34. The Ki-67 PI was evaluated by a double immunostaining technique using the monoclonal antibodies MIB-1 and CD38. Results: MVD, Ki-67 PI, bone marrow infiltration, and CRP were significantly higher in pretreatment patients than in controls and decreased in patients achieving plateau phase. MVD significantly correlated with Ki-67 PI and infiltration, and Ki-67 correlated with infiltration. Conclusion: In multiple myeloma, apart from being a marker of proliferative activity, Ki-67 is also associated with bone marrow angiogenesis and tumour burden.

Interleukin-18 in multiple myeloma patients: serum levels in relation to response to treatment and survival

Interleukin-18 (IL-18) plays a role in the hosts response to tumours and angiogenesis. We determined serum levels of IL-18, vascular endothelial growth factor (VEGF), angiogenin (ANG), tumor necrosis factor (TNF-alpha) and CRP in 65 newly diagnosed myeloma patients. IL-18, VEGF, angiogenin, TNF-alpha and CRP were significantly higher at stage III in comparison to stages II and I. These cytokines (measured in 27 patients) significantly decreased after treatment. In survival analysis, higher levels of IL-18 were associated with a poorer prognosis. We conclude that increased serum IL-18 in myeloma patients correlates with advanced disease, increased levels of angiogenic cytokines and worse survival.

Serum Evaluation of Angiogenic Cytokines Basic Fibroblast Growth Factor, Hepatocyte Growth Factor and TNF-ALPHA in Patients with Myelodysplastic Syndromes: Correlation with Bone Marrow Microvascular Density:

Recent studies have documented that angiogenesis plays a significant role in haematological malignancies, including mylodysplastic syndromes (MDS). Basic fibroblast growth factor (b-FGF), Hepatocyte growth factor (HGF) and Tumor necrosis factor-α (TNF-α) are multifunctional cytokines that potently stimulate angiogenesis. The aim of the present study was to evaluate the microvascular density (MVD) and the serum levels of these angiogenic factors in patients with myelodysplastic syndromes (MDS). In 61 patients with MDS, MVD was measured in bone marrow biopsies and b-FGF, HGF and TNF-α were determined in the serum of the same patients by enzyme-linked immunosorbent assay (ELISA). Serum levels of b-FGF, HGF and TNF-α as well as MVD in the bone marrow were increased in MDS patients compared to healthy controls (p<0.0001). Levels of b-FGF, HGF and TNF-α were also significantly higher in high-risk for leukemic transformation MDS than in low-risk (p<0.0001). Significant differences were also found regarding MVD in high and low risk patients (p<0.001). Both b-FGF and HGF levels were significant predictors of survival (p<0.0005, log-rank test). The present study showed that serum levels of b-FGF, HGF and TNF-α are significantly increased and dependent on the severity of MDS suggesting that the determination of these parameters may offer considerable information regarding disease progression and prognosis.

Angiogenesis-related growth factors and cytokines in the serum of patients with B non-Hodgkin lymphoma; relation to clinical features and response to treatment

Increased angiogenesis has been shown to be a feature of non‐Hodgkin lymphomas (NHL). In the current study, the pretreatment levels of circulating molecules related to angiogenesis were determined in 49 B‐cell NHL patients and correlated with histological grade, disease stage and prognostic score. In 25 patients, the same molecules were defined after standard treatment. Vascular endothelial growth factor (VEGF), angiogenin, interleukin‐2 (IL‐2), IL‐6, IL‐8 and IL‐16 were measured. Increased levels of VEGF, IL‐6 and IL‐8 were found in the whole group of untreated patients in comparison with normal controls (P < 0.05), whereas, IL‐2 was higher in the subgroup of indolent NHL. Overall, there was no significant decrease in the levels of these molecules after treatment. However, by stratification into group of responders vs. non‐responders pretreatment IL‐8 was significantly increased whereas IL‐16 was decreased in the subgroup of complete responders. According to the REAL classification IL‐2 was higher in the low risk compared with intermediate plus high‐risk group. There was no association with disease stage or the International Prognostic Score. Both indolent and aggressive B cell lymphomas have increased production of angiogenic mediators and cytokines with IL‐8 and IL‐16 potentially reflecting the response to treatment.

Role of platelet-derived growth factor-AB in tumour growth and angiogenesis in relation with other angiogenic cytokines in multiple myeloma

Angiogenesis is a complex process essential for the growth, invasion, and metastasis of various malignant tumours, including multiple myeloma (MM). Various angiogenic cytokines have been implicated in the angiogenic process. Among them, platelet‐derived growth factor‐AB (PDGF‐AB) has been reported to be a potent stimulator of angiogenesis in many solid tumours and haematological malignancies, including MM. The aim of the study was to investigate the relationship between PDGF‐AB, microvascular density (MVD), and various angiogenic cytokines, such as basic fibroblast growth factor (b‐FGF), angiogenin (ANG), and interleukin‐6 (IL‐6), in MM patients. Forty‐seven MM patients before treatment, 22 of whom were in plateau phase, were studied. We determined the serum levels of the aforementioned cytokines and MVD in bone marrow biopsies before and after treatment. Mean serum values of PDGF‐AB, b‐FGF, ANG, and MVD were significantly higher in patients compared with controls and with increasing disease stage. Significant positive correlations were observed between serum PDGF‐AB, ANG, and IL‐6 levels and MVD. Furthermore, we found significant positive correlations between PDGF‐AB and b‐FGF, IL‐6, ANG, and β2 microglobulin. We also found that patients with high MVD had statistically significantly higher serum levels of PDGF‐AB when a median MVD value of 7.7 was used as the cutoff point. Furthermore, a significant difference was found in serum levels of PDGF‐AB between pre‐ and post‐treatment patients. Finally, survival time was significantly higher in the low MVD group versus the high MVD group (76 vs 51 months). Our results showed that there is a strong positive correlation between PDGF‐AB and the studied angiogenic cytokines and MVD. It seems that PDGF‐AB plays a role in the complex network of cytokines inducing bone marrow neovascularization in patients with MM. Copyright

Serum BAFF levels are related to angiogenesis and prognosis in patients with multiple myeloma

B-cell activating factor (BAFF) is a B-cell growth factor. We measured its serum levels and correlated them with parameters of disease activity, as serum levels of tumor necrosis factor-α and lactate dehydrogenase, bone marrow microvascular density and proliferating cell nuclear antigen expression, in 50 myeloma patients, in 22 of them in plateau phase and in 20 controls. All of them were higher in patients and in advanced disease while reduced in plateau phase. BAFF correlated with all the above markers. Higher BAFF levels predicted a shorter survival, suggesting an important prognostic marker and a possible therapeutic target in myeloma.

Serum levels of leptin in multiple myeloma patients and its relation to angiogenic and inflammatory cytokines.

BACKGROUND Leptin, apart from the regulation of food intake, has been implicated in hematopoiesis, the immune response and angiogenesis. Leptin has been found to be decreased in various hematological malignancies. In the present study leptin was measured in multiple myeloma (MM) patients before and after treatment and correlated with other angiogenic molecules and markers of disease activity. METHODS Serum leptin, vascular endothelial growth factor (VEGF), basic fibroblast growth factor (b-FGF), interleukin-1 beta (IL-1beta), beta 2 microglobulin (beta2M) and C-reactive protein (CRP) were measured in 62 newly diagnosed MM patients, 22 of whom obtaining disease stabilization after treatment. The same parameters were measured in 20 healthy controls. Disease stage was defined according to the Durie-Salmon criteria. RESULTS Leptin, VEGF, b-FGF, IL-1beta, and beta2M were significantly higher in newly diagnosed MM patients than in controls (p<0.05). VEGF, b-FGF, IL-1beta, beta2M, CRP but not leptin increased with advancing stage of disease (p<0.01). All parameters decreased significantly following treatment (p<0.001). Although IL-1beta correlated positively with VEGF, beta2M, b-FGF and CRP, leptin did not correlate with any of the measured parameters. CONCLUSION Leptin serum levels do not reflect disease severity in MM. However, there seems to be a decrease in leptin following treatment, which may be associated with an alteration in the metabolic state or the chemokine milieu.

Interleukin-10 Induces Both Plasma Cell Proliferation and Angiogenesis in Multiple Myeloma.

In multiple myeloma the angiogenic process is enhanced by various mediators. Among them interleukin-10 (IL-10), secreted mainly by myeloma-associated macrophages seems to participate in myeloma progression with variable manners. The aim of the study was to measure serum levels of IL-10 in various stages of MM patients and to correlate them with various angiogenic cytokines, such as vascular endothelial growth factor and angiopoietin-2 and with known proliferation parameters, such as serum levels of B-cell activating factor and bone marrow infiltration by myeloma plasma cells, in order to explore their clinical significance. We measured serum levels of the above parameters by ELISA in 54 newly diagnosed MM patients. All of them were higher in MM patients and were increasing in parallel with disease progression. Furthermore, IL-10 correlated positively with both angiogenic cytokines and proliferation markers. This correlation of IL-10 with both angiogenic cytokines and markers of disease activity implicates that they all have an important role in MM pathogenesis and progression.

Monitoring serum levels ELR+ CXC chemokines and the relationship between microvessel density and angiogenic growth factors in multiple myeloma

BACKGROUND The ELR+ CXC chemokines are important mediators of tumorigenesis, related to their angiogenic properties. Angiogenesis appears to be a prominent feature in the progression of multiple myeloma (MM). CXC chemokines have four highly conserved cysteine amino acid residues, with the first two cysteine molecules separated by a single amino acid. The angiogenic potential of this group is determined by the presence of three amino acid residues (Glu-Leu-Arg: the ELR motif) preceding the first cysteine amino acid, in the NH2 terminus. AIMS The purpose of this study was to determine serum concentrations of angiogenesis-related chemokines ELR+ motif, such as interleukin-8 (IL-8), epithelial neutrophil activating protein-78 (ENA-78) and growth-related gene alpha (GRO-α), as well the bone marrow microvascular density (MVD) in patients with MM at diagnosis and after treatment, in plateau phase. We also evaluated the relationship among them with other known growth factors involved in angiogenesis. METHODS Serum levels of the ELR+ CXC chemokines: IL-8, ENA-78 and GRO-α as well as of the angiogenic factors: hepatocyte growth factor (HGF), vascular endothelial growth factor (VEGF) and tumor necrosis factor-α (TNF-α) were determined in 63 newly diagnosed MM patients, in 30 in plateau phase and in 20 healthy controls. Serum measurements of them were performed with commercially available kits for ELISA. Bone marrow biopsies were performed before and after treatment, in plateau phase, in order to determine MVD by staining vessels with anti-CD31. RESULTS Serum concentrations of IL-8, ENA-78, GRO-α and TNF-α were significantly higher in the group of MM patients (44.5±25.3, 765±572.1, 186.5±129.1 and 4.2±2.8 pg/ml, respectively) in comparison to control group (27.3±6.4, 335.1±268.6, 112.5±76.1 and 1.3±0.8 pg/ml) (p<0.02 for GRO-α, p<0.001 for other cases). We also found that untreated patients had higher levels of IL-8, ENA-78, GRO-α than post treatment patients, but statistical significant difference was found only for IL-8 (48.36±30.93 pg/ml vs. 35.05±19.77 pg/ml, p<0.001). Furthermore IL-8, GRO-α, TNF-α, HGF and VEGF were significantly higher with increasing disease stage (p<0.001 in all cases). ENA-78 serum levels were higher in stage III than in stage I and II, but without statistical significance. Additionally we correlated each proinflammatory cytokine with well known angiogenic factors such as HGF, VEGF and TNF-α. A positive correlation was found between serum HGF and IL-8 and GRO-α (r=0.316 p<0.01, r=0.297 p<0.02, respectively). Similarly serum VEGF correlated with ENA-78 and GRO-α (r=0.323 p<0.01, r=0.469 p<0.001, respectively). In the pretreatment group of patients a positive correlation between bone marrow MVD and serum levels of GRO-α was found (r=0.304 p<0.01). There was a difference in survival times between patients with higher than median versus low IL-8, ENA-78 and GRO-α levels, but the differences could not reach statistical significance in either case. CONCLUSIONS These findings support the hypothesis that ELR+ motif CXC chemokines, such as IL-8, ENA-78 and GRO-α correlate with angiogenic growth factors and may play a role in the progression of MM. Further studies are needed to determine their prognostic and predictive significance.