Correlation of Mast Cell Density With Angiogenic Cytokines in Patients With Active Multiple Myeloma.

PURPOSE: The aim of the study is to estimate whether bone marrow mast cell density (MCD) in multiple myeloma (MM) correlates with circulating levels of various angiogenic factors. METHODS: In 70 patients with newly diagnosed active MM, we measured MCD using immunohistochemical stain for tryptase and serum levels of matrix metalloproteinase 9 (MMP-9), angiopoietin 2 (ANGIOP-2), and angiogenin (ANG) with ELISA. FINDINGS: Levels of MCD, ANGIOP-2, and ANG were significantly higher in MM patients compared with the control group. The MMP-9 level was higher in MM patients compared with the control group but without statistical significance. All values were increasing in parallel with clinical stages. Furthermore, MCD correlated positively with MMP-9, ANGIOP-2, and ANG. IMPLICATIONS: MCs participate in the angiogenic processes of MM, with complex implicated mechanisms. This interplay between MCs and the other participants favors angiogenesis and MM growth.

High density of tryptase-positive mast cells in patients with multiple myeloma: correlation with parameters of disease activity.

Multiple myeloma (MM) is a plasma cell neoplasm characterized by bone marrow infiltration from malignant plasma cells. Mast cells play an important role in inflammation and angiogenesis in malignant diseases. The aim of the study was to evaluate the mast cell density in bone marrow of untreated MM patients with markers of disease activity such as serum interleukin-6 (IL-6), B2M, and C-reactive protein (CRP), the grade of bone marrow infiltration, and the levels of produced paraprotein. We studied 86 newly diagnosed MM patients (46 males, 40 females, mean age 59 ± 13.7 years). Thirty of them reached plateau phase after chemotherapy and 20 healthy volunteers. According to the criteria of International Staging System (ISS) staging system, 23 patients had stage I, 30 had stage II, and 33 had stage III. The serum concentrations of CRP, B2M, and IL-6, and the mast cell density (MCD) values were significantly higher in MM patients' group (1.6 ± 1.8, 4.3 ± 2.9, 7.1 ± 5.1, and 9 ± 4.8), in comparison with those found in control group (0.4 ± 0.1, 1.5 ± 0.6, 1.1 ± 0.5, and 1.9 ± 0.7; p < 0.001 in all the cases). Significant differences were found between the grade of infiltration in bone marrow, and the paraprotein values in patients' serum before and after chemotherapy. Furthermore, there was a significant correlation between the MCD values and the prognostic markers CRP (r = 0.452, p < 0.0001), IL-6 (r = 0.475, p < 0.0001), bone marrow infiltration (r = 0.333, p < 0.0002), and serum paraprotein levels(r = 0.221, p < 0.04). High MCD values strengthen the hypothesis that mast cells participate in the pathogenesis of disease progression and may be used as an indicator of the disease activity.

Mast cells influence the proliferation rate of myeloma plasma cells.

In multiple myeloma (MM), mast cells (MCs) modify bone marrow microenvironment. In order to estimate whether MC density (MCD) in active MM bone marrows relates to the proliferative activity of plasma cells, we estimated in 42 patients MCD, microvascular density (MVD), and the Ki-67 proliferation index (PI) (immunohistochemical expression of tryptase, CD31, and Ki-67). MCD correlated with Ki-67 PI (p < .001), suggesting the important participation of MCs in MM biology and growth; MCs enhance angiogenesis and produce cytokines with growth effects on myeloma cells. Therefore, MCs could be valuable targets for therapeutic interventions.

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.

Immunohistochemical expression of endoglin offers a reliable estimation of bone marrow neoangiogenesis in multiple myeloma.

PURPOSE: The aim of the present study was to evaluate CD105 tissue marker in the bone marrow (BM) of multiple myeloma (MM) patients. CD105 was evaluated using immunohistochemical method. An effort was made to correlate this marker with BM microvascular density (MVD) along with other known markers of angiogenesis in order to evaluate its clinical significance. METHODS: BM MVD was estimated by CD31. CD105 in BM was estimated by immunohistochemical method in 54 newly diagnosed patients with MM. Circulating levels of known angiogenic factors such as basic fibroblast growth factor (b-FGF) and soluble CD105 (sCD105) were measured by ELISA in the same group of patients. All these factors were also measured in 20 age- and sex-matched healthy controls. RESULTS: We found that CD105 MVD, along with the expected CD31 MVD, and serum levels of sCD105 and bFGF were increased, also in parallel with disease stage, and all were decreased after effective treatment. Moreover, CD105 MVD correlated with all the aforementioned markers of angiogenesis. CONCLUSIONS: Our results indicated that CD105 MVD is following the behavior of CD31 MVD in MM, suggesting being a valid marker of BM neoangiogenesis in MM. Its prognostic impact remains to be proven.

The Impact of Mast Cell Density on the Progression of Bone Disease in Multiple Myeloma Patients.

BACKGROUND: Osteolytic bone disease is a major hallmark in multiple myeloma (MM) progression and affects many patients. Several inflammatory cells are involved in MM progression. Among them, mast cells (MCs) accumulated in the bone marrow (BM) microenvironment are known to play an important role in the mechanism of neovascularization. METHODS: In 52 newly diagnosed active MM patients, we measured BM MC density (MCD) using an immunohistochemical stain for tryptase, serum levels of matrix metalloproteinase-9 (MMP-9) and receptor activator of nuclear factor x03BA;B ligand (RANKL) by a solid-phase sandwich enzyme-linked immunosorbent assay, along with urine levels of N-terminal propeptide of procollagen type I (Ntx) by a competitive inhibition enzyme-linked immunosorbent assay, in various clinical stages and skeletal grades. RESULTS: MCD, RANKL and Ntx were higher in MM patients. All values increased in association with both the clinical stage and skeletal grade. Furthermore, MCD correlated positively with MMP-9, RANKL and Ntx. CONCLUSIONS: Our data suggest that MCs may contribute to osteolytic processes during MM progression. Although the major role of MCs in tumor progression is to enhance angiogenesis, it seems that they may affect MM bone disease and may secrete a plethora of mediators that may directly and indirectly have an impact on osteolysis.

Clinical significance of interleukin-22 in multiple myeloma.

OBJECTIVE: Interleukin-22 (IL-22) is a cytokine participating in many aspects of inflammation. Multiple myeloma (MM) is a malignant disease of plasma cells with characteristic immune deregulation. We estimated serum levels of IL-22 in MM patients, both in activity and remission, in order to apprehend its possible participation in MM biology. METHODS: We measured serum levels of IL-22 along with beta-2 microglobulin (B2M), paraprotein, and interleukin-1beta (IL-1beta), as well as degree of bone marrow infiltration, in 51 patients with active MM and in 22 of them in remission. RESULTS: We found that IL-22 was higher in active MM patients, compared to both controls and patients in remission, and also in patients in remission compared to controls. Moreover, IL-22 was increasing in parallel with the disease stage and also correlated with B2M, IL1-beta, and degree of infiltration. DISCUSSION: We suggest that the elevated levels of IL-22 in active MM patients, in parallel with disease activity, and in positive correlation with IL-1beta, may represent the inflammatory element of the disease. This increased occurrence of IL-22 may enhance myeloma proliferation and growth, and moreover, may participate in the mechanisms of immune deregulation.

Prognostic impact of angiopoietin-2 in multiple myeloma.

PURPOSE: Angiogenesis is an essential process for the expansion of multiple myeloma (MM). Angiopoietin-2 (Ang-2), Ang-1 and their receptor possess important roles in this procedure. The aim of the study was to measure serum levels of Ang-2 along with known markers of angiogenesis and to estimate their prognostic impact on the survival. METHODS: Bone marrow microvascular density (MVD), estimated by CD31, and circulating levels of known angiogenic factors Ang-2, interleukin-6, soluble CD105 and platelet-derived growth factor-AB, measured by ELISA, were measured in 77 newly diagnosed patients with active MM and in 57 of them who responded to chemotherapy. RESULTS: All measured parameters were increased in MM patients, were also increasing in advanced disease and decreased after effective treatment. Ang-2 correlated positively with the other angiogenic factors and MVD. Moreover, Ang-2 values above the median were accompanied by worse survival. CONCLUSION: Ang-2 correlates strongly with the angiogenic process and its serum levels are importantly prognostic for survival, highlighting the role of angiopoietins pathway in the biology of MM.

Circulating levels of soluble Fas ligand reflect disease progression in multiple myeloma.

Multiple myeloma (MM) plasma cells are apoptosis resistant. The system of Fas with its ligand (Fas-L) participates actively in the extrinsic apoptotic system. In oncology, its role is controversial, since it has been reported both to suppress and promote tumor growth. The aim of this study was to measure serum levels of soluble Fas-L (sFas-L) in patients with active MM and to correlate them with markers of disease activity. We studied 57 patients with active MM, along with 22 healthy controls. We measured serum levels of sFas-L and interleukin-6 (IL-6) by enzyme-linked immunosorbent assays, as well of beta-2 microglobulin (B2M), C-reactive protein (CRP) and lactate dehydrogenase (LDH). We also measured the degree of bone marrow infiltration. All parameters were increased in patients, compared with controls (p < 0.001 for all cases) and also in parallel with disease stage (p < 0.001 for all cases). Positive correlations were noted between serum levels of sFas-L with IL-6, infiltration (p < 0.001 for both cases) and LDH (p < 0.04), but not with CRP and B2M. We suggest that the system of Fas/Fas-L participates actively in MM progression in a complex manner and that serum levels of sFas-L may reflect disease progression. Further studies are needed to determine its usefulness as a marker of disease activity.

Increased serum levels of MIP-1alpha correlate with bone disease and angiogenic cytokines in patients with multiple myeloma.

Many cytokines possess variable roles in the pathogenesis of multiple myeloma. Macrophage inflammatory protein-1alpha (MIP-1alpha) is an osteoclast-activating factor with a major role in myeloma bone disease. The aim of the study was to examine its participation in the angiogenic process of the disease. We measured, by enzyme-linked immunosorbent assays, its serum levels in 56 newly diagnosed myeloma patients, in several skeletal grades and stages of the disease and in 25 healthy controls. Concurrently, we measured serum levels of the angiogenic cytokines basic-fibroblast growth factor, hepatocyte growth factor and interleukin-18. All the above cytokines were higher in myeloma patients (p < 0.001 for all cases) and were increasing in parallel with disease stage (p < 0.001 for all cases) and skeletal grade (p < 0.04 for MIP-1alpha and p < 0.001 for the other cases). Moreover, positive correlations between MIP-1alpha and all the angiogenic cytokines were noted (p < 0.001 for all cases). MIP-1alpha seems to be a predominant factor responsible for the enhancement of bone resorption and increased angiogenesis. The positive correlation between MIP-1alpha and the angiogenic chemoattractants supports the involvement of these factors in the biology of myeloma cell growth. Moreover, they could be used as possible therapeutic targets as well as markers of disease activity.

Positive correlation between bone marrow mast cell density and ISS prognostic index in patients with multiple myeloma.

We evaluated mast cell density (MCD) in myeloma bone marrow biopsies and correlated it with stage of disease and markers of angiogenesis. Fifty-three untreated myeloma patients and 28 of them responded to therapy were studied. Mast cells were highlighted using immunohistochemical stain for tryptase. Angiogenesis was evaluated measuring microvascular density and serum levels of basic-fibroblast growth factor and tumor necrosis factor-alpha. MCD was higher in untreated patients, compared to healthy population and responders. Significant association was found between MCD with angiogenesis and clinical stage of disease, suggesting that mast cells could be used as target for myeloma treatment.

Relationship between circulating BAFF serum levels with proliferating markers in patients with multiple myeloma.

In multiple myeloma, there are many factors influencing the growth of the malignant clone in direct and indirect manners. BAFF is a growth factor for myeloma cells. The aim of the study was to measure its circulating levels in 54 pretreatment patients, along with serum levels of other proliferation markers, such as interleukins-6, -10, and -15, CRP, and beta-2 microglobulin, as well as bone marrow plasma cell infiltration and expression of Ki-67 PI, in various stages of the disease and after effective treatment in 28 of them. Serum levels of the previously mentioned factors were measured by ELISA, whereas bone marrow plasma cell infiltration and Ki-67 expression were estimated immunohistochemically. All measured parameters were higher in pretreated myeloma patients compared to healthy population and were also increasing with the progression of the disease. They all also decreased after effective therapy. Furthermore, all pretreatment values correlated to each other. BAFF seems to be an important growth factor for myeloma plasma cells. Measuring its serum levels, along with the previously mentioned cytokines, may provide important information regarding the degree of myeloma cells' proliferation. Therefore, they all could be used as markers of proliferation and disease activity.

Serum levels of angiopoietin-2 are associated with the growth of multiple myeloma.

Angiopoietins and their receptor, Tie-2, have crucial role in angiogenesis. We measured serum levels of angiopoietin-2 (Ang-2), soluble Tie-2, and factors of burden and prognosis in myeloma (LDH, CRP, beta-2 microglobulin, and interleukin-6) in 55 newly diagnosed patients, with 30 of them in plateau phase, in order to note correlations among them. Levels of Ang-2 were higher in patients in advanced stage of disease, decreased in plateau phase, and correlated with all other factors. Circulating Ang-2 in myeloma patients significantly correlated to factors of disease burden and prognosis, and therefore measuring its levels may be important for the valuation of the disease.

The significance of non correlation between interleukin-8 serum levels with bone marrow microvascular density in patients with myeloma multiple.

In multiple myeloma (MM), angiogenesis plays a substantial role in disease progression. Interleukin-8 (IL-8), a pro-inflammatory chemokine with potent pro-angiogenic properties, has been implicated in the pathophysiology of MM. The aim of the study is to measure serum levels of IL-8 in MM patients and to correlate them with markers of angiogenesis, such as circulating levels of platelet derived growth factor-AB (PDGF-AB) and angiogenin (Ang), and bone marrow microvascular density (MVD). Fifty-three newly diagnosed MM patients, 23 of them, who reached plateau phase after effective treatment and 20 healthy controls, were studied. Serum levels of PDGF-AB, Ang and IL-8 were measured by ELISA, whereas bone marrow MVD was estimated by immunohistochemical staining of vessels with anti-CD31. All measured parameters were higher in MM patients compared to controls and in increased disease stages. They all also significantly decreased in plateau phase. IL-8 correlated positively with Ang and PDGF-AB, but not with MVD. The circulating levels of IL-8, PDGF-AB and Ang are elevated in patients with MM. The lack of correlation between IL-8 with MVD suggests that its levels represent the inflammatory element of MM disease and the participation in angiogenesis process is rather complex with multifactorial mechanisms.

The relationship between soluble receptor of interleukin-6 with angiogenic cytokines and proliferation markers in multiple myeloma.

Soluble interleukin-6 receptor (sIL-6R) is part of IL-6 receptor that may stimulate cells that do not express the whole molecule. It may enhance myeloma cell proliferation and furthermore angiogenesis. The aim of the study was to evaluate the clinical significance and the relationship between serum levels of sIL-6R, with various stimulators of angiogenesis, such as hepatocyte growth factor (HGF) and interleukin-18 (IL-18) and with markers of proliferation, such as beta-2 microglobulin (B2M) levels and plasma cell Ki-67 proliferation index in the bone marrow, in patients with multiple myeloma (MM). We studied 45 newly diagnosed MM patients. Serum levels of sIL-6R, HGF, IL-18, and B2M and Ki-67 proliferation index (Ki-67 PI) in bone marrow's plasma cells were determined. The mean concentrations of sIL-6R, HGF, IL-18, and B2M and the value of Ki-67 were significantly higher in the patients compared to controls and with increasing disease stage. sIL-6R was strongly positively correlated with HGF, IL-18, B2M, and Ki-67 PI. There is a positive correlation between plasma cell growth, as determined by Ki-67 PI, and different angiogenic cytokines, such as HGF and IL-18, with sIL-6R. This relationship suggests the significant role of these cytokines in the proliferation and disease activity in MM patients.

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 ß2 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.

Serum interleukin-17 and its relationship to angiogenic factors in multiple myeloma.

BACKGROUND: Interleukin-17 (IL-17) is a CD4 T-cell-derived mediator of angiogenesis that stimulates vascular endothelial cell migration and regulates the production of a variety of proangiogenic factors, such as tumor necrosis factor-alpha (TNF-alpha) and vascular endothelial cell growth factor (VEGF). Angiogenesis is implicated in the progression of multiple myeloma (MM). METHODS: We measured serum levels of IL-17, TNF-alpha, and VEGF, as well as microvessel density (MVD) in 40 untreated MM patients. RESULTS: Levels of IL-17 in the sera of patients with MM were higher than those in matched controls; however, the difference did not reach statistical significance. Serum levels of both TNF-alpha and VEGF in MM patients were significantly higher than those in controls (p<0.001 in both instances). Levels of IL-17 in MM patients, both stage II and stage III, were significantly higher than those of stage I patients (p=0.001 and p<0.001, respectively). Similarly, higher values of VEGF (p<0.001), TNF-alpha (p<0.001), and MVD (p<0.035) were associated with advanced disease stage. Serum values of IL-17 in MM patients correlated positively not only with VEGF (Spearman's rho=0.606) and TNF-alpha (r=0.552; p<0.001 in both instances), but also with MVD (r=0.385, p=0.014). In addition, a positive correlation was found between serum values of VEGF and TNF-alpha (r=0.657, p<0.001), MVD and VEGF (r=0.353, p=0.026), and between MVD and TNF-alpha (r=0.506, p=0.001) in MM patients. CONCLUSION: These results suggest that IL-17 plays a role in the promotion of angiogenesis and associated disease progression in MM.

Relation between bone marrow angiogenesis and serum levels of angiogenin in patients with myelodysplastic syndromes.

Angiogenesis is implicated in the progression of myelodysplastic syndromes (MDS). Bone marrow microvascular density (MVD), serum angiogenin (ANG) and interleukin 6 (IL-6) were measured in 67 patients with untreated MDS. MVD, ANG and IL-6 were significantly higher in the patient group as a whole when compared to controls (P < 0.01). MVD and ANG were significantly higher in subtypes with a high-risk for leukemic transformation (RAEB, RAEB-t and CMML) than in low-risk subtypes (RA and RARS) (P < 0.01). In the MDS group, a positive correlation was found between ANG and IL-6 (P < 0.001) and also between MVD and IL-6 (P < 0.05). Using multivariate analysis, only IL-6 displayed independent prognostic value and was inversely related to MDS survival.

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

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.