In vivo modulation of angiogenesis by beta 2 glycoprotein I.

Beta 2 glycoprotein I (ß2GPI) is the major auto antigen in the antiphospholipid syndrome but also interacts with fibrinolytic and angiogenic proteins. The aim of this study was to examine the angiogenic potential of ß2GPI in vivo in ß2GPI deficient mice utilizing angiogenic assays. ß2GPI deficient mice show increased microvessel formation in comparison to ß2GPI replete controls when injected with growth factor free-matrigel implants. However, microvessel formation in matrigel plugs of ß2GPI deficient mice was less than in ß2GPI replete mice when basic fibroblast growth factor (bFGF) was included in the matrigel. Hemoglobin content was higher in vascular endothelial growth factor (VEGF) containing-matrigel plugs in the ß2GPI deficient mouse demonstrating that the lack of ß2GPI led to increased extravasation by VEGF. Melanoma B16F10 tumour growth was enhanced in ß2GPI deficient mice. Melanoma microvessel density was increased in ß2GPI deficient mice but the proliferation rate of tumour cells (determined by Ki67 immunohistochemistry) was unaffected by the presence or absence of ß2GPI. Subcutaneous delivery of native human ß2GPI by the ALZET osmotic pump did not affect melanoma tumour growth in ß2GPI deficient mice. We conclude that the in vivo unopposed action of ß2GPI is anti-angiogenic however this function is modified in the presence of a strong angiogenic stimulus into stabilization of vessel formation. Although the presence of ß2GPI attenuates vessel sprouting in certain tumours, no survival benefit is conferred to tumour bearing animals. This does not preclude the potential benefit of modified or fragments of ß2GPI in anti-angiogenesis research.

Quantification of NETs-associated markers by flow cytometry and serum assays in patients with thrombosis and sepsis.

INTRODUCTION: Neutrophil extracellular traps (NETs) are networks of extracellular fibres produced from neutrophil DNA with a pathogenic role in infection, thrombosis and other conditions. Reliable assays for measuring NETs are desirable as novel treatments targeting NETs are being explored for the treatment of these conditions. We compare a whole blood flow cytometry method with serum assays to measure NETs-associated markers in patients with sepsis and thrombosis. METHODS: Patients with deep venous thrombosis (n = 25), sepsis (n = 21) and healthy controls (n = 23) were included in the study. Neutrophil surface NETs markers were determined by flow cytometry on whole blood samples by gating of neutrophils stained for surface citrullinated histone (H3cit) and myeloperoxidase (MPO). Serum double-stranded (ds) DNA, MPO, myeloid-related protein, nucleosomes, DNAse, elastase, human high-mobility group box 1 and MPO-DNA complexes were quantified as circulating markers of NETs. RESULTS: Neutrophil NETs markers by flow cytometry and serum NETs markers were significantly higher in patients with thrombosis and sepsis compared with healthy controls. Neutrophil NETs markers significantly correlated with the serum marker dsDNA. CONCLUSION: Flow cytometry detection of neutrophil NETs markers is feasible in whole blood and correlates with serum markers of NETs. We propose the flow cytometry detection of MPO/H3cit positive neutrophils and serum dsDNA as simple methods to quantify cellular and extracellular NET markers in patients with thrombosis and sepsis.

Protein disulfide isomerase regulation by nitric oxide maintains vascular quiescence and controls thrombus formation.

Essentials Nitric oxide synthesis controls protein disulfide isomerase (PDI) function. Nitric oxide (NO) modulation of PDI controls endothelial thrombogenicity. S-nitrosylated PDI inhibits platelet function and thrombosis. Nitric oxide maintains vascular quiescence in part through inhibition of PDI. SUMMARY: Background Protein disulfide isomerase (PDI) plays an essential role in thrombus formation, and PDI inhibition is being evaluated clinically as a novel anticoagulant strategy. However, little is known about the regulation of PDI in the vasculature. Thiols within the catalytic motif of PDI are essential for its role in thrombosis. These same thiols bind nitric oxide (NO), which is a potent regulator of vessel function. To determine whether regulation of PDI represents a mechanism by which NO controls vascular quiescence, we evaluated the effect of NO on PDI function in endothelial cells and platelets, and thrombus formation in vivo. Aim To assess the effect of S-nitrosylation on the regulation of PDI and other thiol isomerases in the vasculature. Methods and results The role of endogenous NO in PDI activity was evaluated by incubating endothelium with an NO scavenger, which resulted in exposure of free thiols, increased thiol isomerase activity, and enhanced thrombin generation on the cell membrane. Conversely, exposure of endothelium to NO+ carriers or elevation of endogenous NO levels by induction of NO synthesis resulted in S-nitrosylation of PDI and decreased surface thiol reductase activity. S-nitrosylation of platelet PDI inhibited its reductase activity, and S-nitrosylated PDI interfered with platelet aggregation, α-granule release, and thrombin generation on platelets. S-nitrosylated PDI also blocked laser-induced thrombus formation when infused into mice. S-nitrosylated ERp5 and ERp57 were found to have similar inhibitory activity. Conclusions These studies identify NO as a critical regulator of vascular PDI, and show that regulation of PDI function is an important mechanism by which NO maintains vascular quiescence.

Angiogenic molecules in Hodgkin's disease: results from sequential serum analysis.

Increased angiogenic activity has been demonstrated in lymphoproliferative diseases including Hodgkin's disease. In the current study, the levels of circulating angiogenic molecules in 60 Hodgkin's patients were determined prior to and after treatment and correlated to disease stage and prognostic score. Hepatocyte growth factor (HGF), vascular endothelial growth factor (VEGF), interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-alpha) were increased in Hodgkin's patients in comparison to healthy controls (p<0.001). Angiogenin and angiopoietin-2 levels did not differ from controls. HGF, VEGF, TNF-alpha and angiogenin decreased significantly in Hodgkin's patients after standard treatment (p<0.001 for HGF, p<0.05 for VEGF, TNF-alpha and angiogenin). Furthermore, HGF and TNF-alpha increased with advancing stage of disease (p<0.05). HGF and VEGF correlated significantly with IL-6 (r=0.56, p<0.0005 and r=0.57, p<0.001 respectively). In conclusion, Hodgkin's disease displays an angiogenic activity as depicted by the increased serum levels of a number of angiogenic cytokines. HGF seems to be the prominent molecule in Hodgkin's disease, which may be used to monitor the disease status and the response to treatment.

Serum evaluation of angiogenic cytokine 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-alpha (TNF-alpha) 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-alpha were determined in the serum of the same patients by enzyme-linked immunosorbent assay (ELISA). Serum levels of b-FGF, HGF and TNF-alpha 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-alpha 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-alpha 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.

The assessment of proliferating cell nuclear antigen immunostaining in myelodysplastic syndromes and its prognostic significance.

Several prognostic factors for patients with myelodysplastic syndromes (MDS) have been identified in previous years. In order to determine prognostic factors characterizing haematopoietic cell kinetics, bone marrow proliferative activity and serum TNF-a levels were measured in 51 cases of MDS. Cell proliferation was evaluated by employing a monoclonal antibody directed against the proliferating cell nuclear antigen (PCNA). The PCNA proliferating index (PCNA PI) and serum TNF-a levels showed significant differences between patients with MDS and normal controls (p<0.0001). PCNA PI and serum TNF-a were significantly higher in the high risk for leukemic transformation FAB subgroups (RAEB, RAEB-t and CMML) in comparison to the low risk group (RA and RARS) (p<0.001). PCNA PI and TNF-a also increased with increasing IPSS score (p<0.05). A positive correlation was noted between TNF-a concentrations and PCNA PI (r:0.36, p<0.008). Univariate analysis using the log-rank test showed that a higher PCNA PI was associated with a significantly shorter survival (p<0.001). We conclude that elevated PCNA PI and TNF-a serum levels are increased in high risk myelodysplastic disease and that a high PCNA PI is predictive of a shorter survival in this group of patients.

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 host's 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.

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.

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.

Serum proinflammatory cytokines and its relationship to clinical parameters in lung cancer patients with reactive thrombocytosis.

Proinflammatory cytokines Interleukin-1 beta (IL-1 beta) and Interleukin-6 (IL-6) play a significant role in the pathogenetic processes related to various malignant and inflammatory conditions. Leukocytosis, thrombocytosis and increased acute phase protein levels are part of a systemic inflammatory response. In this study, we measured the concentrations of IL-1 beta, IL-6 and ferritin as well as hemoglobin, lactate dehydrogenase (LDH), C-reactive protein (CRP) and erythrocyte sedimentation rate (ESR) in 23 patients (male 15, female 8, median age 68 years) with lung cancer and reactive thrombocytosis (LCRT), in 27 (male 18, female 9, median age 64 years) with benign inflammatory lung disorder (BILD) and 18 (male 10, female 8, median age 62 years) lung cancer patients with a normal platelet count (LCNP). IL-1 beta levels were significantly higher in the three patient groups in comparison with control subjects (P < 0.001) but without significant difference among the three patient groups. IL-6 was higher in all three patients groups but only in the BILD group it was significantly higher than the control group (P < 0.05). However, no significant difference in IL-6 serum levels was found between the two lung cancer groups. CRP and LDH were significantly higher in the LCRT group in comparison with the other two patient groups (P < 0.01 and 0.001, respectively), while ferritin was higher in both lung cancer groups in comparison with the BILD group (P < 0.001). Our data suggest that in lung cancer patients, reactive thrombocytosis is part of the systemic inflammatory reaction for which IL-1 beta and IL-6 may be intermediate but not independent mediators.

Expression of proliferating cell nuclear antigen (PCNA) in multiple myeloma: its relationship to bone marrow microvessel density and other factors of disease activity.

The expression of proliferating cell nuclear antigen (PCNA) was studied in plasma cells in bone marrow biopsies from patients with multiple myeloma (MM) using a double immunostaining method. In the same samples, microvessel density (MVD), after staining with anti-CD34 antibodies, was determined before and after chemotherapy. The correlation of PCNA expression and MVD with other myeloma parameters (clinical stage, bone marrow plasma cell infiltration and serum interleukin-6 (IL-6)) was also investigated. The study population included 51 newly diagnosed MM patients, 15 patients in plateau phase after treatment and 15 normal controls. Pretreatment mean +/- SE values of PCNA, MVD, plasma cell infiltration and serum IL-6 were significantly higher than post treatment values and controls. Pretreatment PCNA expression correlated significantly with bone marrow MVD (p<0.05) plasma cell infiltration (p<0.01) and IL-6 (p<0.01). These findings show that the proliferative activity of plasma cells is related to the angiogenic activity in the bone marrow of multiple myeloma patients. Both PCNA and MVD correlate with markers of disease activity thus may provide additional information when included in the initial evaluation of myeloma bone marrow biopsies.

Molecular pathophysiology of the antiphospholipid syndrome: the role of oxidative post-translational modification of beta 2 glycoprotein I.

It has been well established that antiphospholipid antibodies and specifically those directed against beta 2 glycoprotein I (ß2GPI) are pathogenic for the development of thrombosis in the antiphospholipid syndrome (APS). Several groups have shown that anti-ß2GPI antibodies, in complex with ß2GPI, elicit effects on blood cells and coagulation-fibrinolysis proteins, which prime the arterial and venous vasculature for the development of thrombosis. However, much less is known about the mechanism initiating the production of autoantibodies against ß2GPI, a physiological abundant protein of blood. In the current review, novel findings are presented regarding the structure and oxidative post-translational modifications of ß2GPI, which trigger the immune response. The majority of circulating ß2GPI exists in a form containing unpaired cysteines (free thiols), which constitutes the reduced form of ß2GPI. The free thiols exposed on ß2GPI are involved in the interaction with platelets and endothelial cells. We propose that this abundant pool of free thiols may serve as an antioxidant reservoir protecting cells or critical molecules from oxidative stress. Oxidation of ß2GPI confers an increase in its immunogenicity through a Th1 immunological mechanism. The clinical significance of these observations is that serum from patients with APS, assessed by a novel ELISA assay, have a significant increase in oxidised ß2GPI. These findings hold promise, not only for the delineation of the role of ß2GPI as an immunological target, but also for the development of improved diagnostic and prognostic assays for APS.

Redox control of ß2-glycoprotein I-von Willebrand factor interaction by thioredoxin-1.

BACKGROUND: ß(2) -Glycoprotein I (ß(2) GPI) is an abundant plasma protein that is closely linked to blood clotting, as it interacts with various protein and cellular components of the coagulation system. However, the role of ß(2) GPI in thrombus formation is unknown. We have recently shown that ß(2) GPI is susceptible to reduction by the thiol oxidoreductases thioredoxin-1 and protein disulfide isomerase, and that reduction of ß(2) GPI can take place on the platelet surface. METHODS: ß(2) GPI, reduced by thioredoxin-1, was labeled with the selective sulfhydryl probe N(a)-(3-maleimidylpropionyl)biocytin and subjected to mass spectrometry to identify the specific cysteines involved in the thiol exchange reaction. Binding assays were used to examine the affinity of reduced ß(2) GPI for von Willebrand factor (VWF) and the effect of reduced ß2GPI on glycoprotein (GP)Ibα binding to VWF. Platelet adhesion to ristocetin-activated VWF was studied in the presence of reduced ß(2) GPI. RESULTS: We demonstrate that the Cys288-Cys326 disulfide in domain V of ß(2) GPI is the predominant disulfide reduced by thioredoxin-1. Reduced ß(2) GPI in vitro displays increased binding to VWF that is dependent on disulfide bond formation. ß(2) GPI reduced by thioredoxin-1, in comparison with non-reduced ß(2) GPI, leads to increased binding of GPIbα to VWF and increased platelet adhesion to activated VWF. CONCLUSIONS: Given the importance of thiol oxidoreductases in thrombus formation, we provide preliminary evidence that the thiol-dependent interaction of ß(2) GPI with VWF may contribute to the redox regulation of platelet adhesion.

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.

Beta2-glycoprotein I inhibits vascular endothelial growth factor and basic fibroblast growth factor induced angiogenesis through its amino terminal domain.

BACKGROUND: Beta-2 glycoprotein I (beta(2)GPI) is a plasma glycoprotein which interacts with various proteins of the coagulation and fibrinolysis system. beta(2)GPI has recently been shown to have anti-angiogenic properties. OBJECTIVES: We undertook this study to investigate the specific domain of beta(2)GPI involved in the anti-angiogenic function and its effect on downstream signaling of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF). METHODS: Various preparations of beta(2)GPI were used on human umbilical vein endothelial cells (HUVECs) in the absence or presence of VEGF and bFGF. The effect on HUVECs' proliferation, migration and tubule formation in Matrigel matrix was investigated. The effect of beta(2)GPI on the mRNA expression of VEGF receptors and phosphorylation of signaling molecules was also studied. RESULTS: beta(2)GPI is shown in this study to be an anti-angiogenic molecule in vitro by inhibiting VEGF and bFGF-induced proliferation, migration and papillary-like tubule formation of HUVECs. This inhibition was achieved by native, proteolytically clipped and domain deletion mutants, domain I-IV (DI-IV) but not domain II-V (DII-V) of beta(2)GPI. Native beta(2)GPI was found to downregulate the expression of the VEGF receptor KDR/Flk-1 on endothelial cells and to block the phosphorylation of VEGF's downstream effector molecules in the MAPK/ERK and PI3K/Akt/GSK3beta pathways. CONCLUSIONS: These results indicate that beta(2)GPI has anti-angiogenic functions which depend on the presence of domain I. This anti-angiogenic activity may have important implications for the therapeutic manipulation of angiogenesis in various disease states.

Effect of different levels of pressure support and proportional assist ventilation on breathing pattern, work of breathing and gas exchange in mechanically ventilated hypercapnic COPD patients with acute respiratory failure.

BACKGROUND: Proportional assist ventilation (PAV) has been shown to maintain better patient-ventilator synchrony than pressure support ventilation (PSV); however, its clinical advantage regarding invasive ventilation of COPD patients has not been clarified. OBJECTIVES: To compare the effect of PAV and PSV on respiratory parameters of hypercapnic COPD patients with acute respiratory failure (ARF). METHODS: Nine intubated hypercapnic COPD patients were placed on the PAV or PSV mode in random sequence. For each mode, four levels (L1-L4) of support were applied. At each level, blood gases, flow, tidal volume (VT), airway pressure (Paw), esophageal pressure (Pes) (n = 7), patient respiratory rate (fp), ventilator rate (fv), missing efforts (ME = fp - fv) were measured. RESULTS: We found increases in ME with increasing levels of PSV but not with PAV. PO2 and VT increased whereas PCO2 decreased significantly with increasing levels of PSV (p < 0.05). With PAV, PCO2 decreased and VT increased significantly only at L4 whereas PO2 increased from L1 to L4. Runaways were observed at L3 and L4 of PAV. The pressure-time product (PTP) was determined for effective and missing breaths. The mean total PTP per minute (of effective plus missing breaths) was 160 +/- 57 cm H2O/s.min in PSV and 194 +/- 60 cm H2O/s.min in PAV. CONCLUSION: We conclude that in COPD patients with hypercapnic ARF, with increasing support, PSV causes the appearance of ME whereas PAV develops runaway phenomena, due to the different patient-ventilator interaction; however, these do not limit the improvement of blood gases with the application of both methods.

Value of Tc-99m sestamibi scintigraphy in the detection of bone lesions in multiple myeloma: comparison with Tc-99m methylene diphosphonate.

Technetium 99m-2-methoxyisobutylisonitrile (Tc-99m MIBI) is a lipophilic agent that accumulates preferentially within living malignant cells due to the higher transmembrane electrical potential as a consequence of the higher metabolic rate than in the surrounding normal cells. It has been effectively used to detect malignant tumors at diagnosis and follow-up and has been reported to be useful in detecting disease lesions in multiple myeloma. We studied 28 consecutive patients with multiple myeloma at diagnosis to determine the value of Tc-99m MIBI in comparison with Tc-99m methylene diphosphonate (MDP), conventional X-rays, computed tomography (CT) scan, and magnetic resonance imaging (MRI). We found 26 patients with obvious osteolytic lesions in X-rays, 22 patients with positive Tc-99m MIBI scans, and 15 patients with positive Tc-99m MDP scans. There was no coincidence of the positive lesions in the two scans, while in two patients the osteolytic areas were positive in the Tc-99m MDP scans, and in one case the osteolytic area was positive in the Tc-99m MIBI scan. The intensity of Tc-99m MIBI scans correlated with disease activity as determined by lactate dehydrogenase (LDH) (p<0.05), C-reactive protein (CRP) (p<0.01), beta2-microglobulin (p<0.05), and serum ferritin (p<0.01). We believe that Tc-99m MIBI scintigraphy can detect bone marrow lesions in myeloma patients that cannot be detected by other imaging methods and that it can be useful especially in solitary myeloma to exclude other involved sites. In addition, it could be a prognostic factor related to disease activity and multidrug resistance. We believe that a multicenter study is needed to evaluate the usefulness of this agent.

Urinary N-telopeptide levels in multiple myeloma patients, correlation with Tc-99m-sestaMIBI scintigraphy and other biochemical markers of disease activity.

Urinary cross-linked N-telopeptide of type I collagen (NTx) has been reported to be a sensitive and specific marker of bone resorption in multiple myeloma (MM). In this study, we measured the levels of NTx in 30 newly diagnosed MM patients and 25 controls. We examined its association with the overall score of skeletal involvement measured by Tc-99m-MIBI scintigraphy and other biochemical markers of bone disease (tumour necrosis factor a (TNF-a), serum calcium and creatinine). We further studied the correlation of NTx with the stage of disease (according to Durie-Salmon criteria) and bone marrow infiltration by plasma cells. High levels of NTx, bone marrow infiltration, TNF-alpha, calcium and creatinine were noted at advanced stages of disease (p < 0.05). NTx and TNF-a were found at significantly higher concentrations in patients with a high overall score (3 and 4) in Tc-99m-sestaMIBI in comparison to a low score (0, 1 and 2; p < 0.05). Positive correlations were found between NTx and TNF-a, as well as between bone infiltration and TNF-a or calcium. In conclusion, NTx is a useful marker for the monitoring of bone resorption in MM and correlates with imaging findings on Tc-99m-sestaMIBI and other biochemical markers of disease activity.

The clinical and prognostic significance of erythrocyte sedimentation rate (ESR), serum interleukin-6 (IL-6) and acute phase protein levels in multiple myeloma.

Interleukin-6 (IL-6) and acute phase proteins are commonly increased in patients with multiple myeloma. Several of these acute phase proteins are believed to predict prognosis and influence survival. We measured interleukin-6 (IL-6), C-reactive protein (CRP), alpha-1-antitrypsin (a1AT), acid alpha-1-glycoprotein (a1AG), haptoglobin (HAP), transferrin (TRF), hemoglobin (Hb), beta-2-microglobulin (beta2M) and erythrocyte sedimentation rate (ESR) in 42 newly diagnosed multiple myeloma patients and 25 normal controls. At the time of blood collection, nine patients were at stage I of disease, 14 at stage II, and 19 at stage III according to the Durie and Salmon myeloma staging system. Mean +/- SD values of IL-6, CRP, a1AT, a1AG, HAP, beta2M, and ESR were significantly higher and Hb significantly lower than those found in the controls. Univariate analysis, using the log-rank test, showed that among the acute phase proteins, serum CRP (P < 0.002), a1AT (P < 0.008) and ESR (P < 0.008) were significantly correlated with survival. However, when a multivariate Cox proportional hazard model was performed, ESR, CRP, a1AT, a1AG and beta2M were identified as independent prognostic factors, while the others were not. We conclude that ESR, a simple and easily performed marker, was found to be an independent prognostic factor for survival in patients with multiple myeloma.

Microsatellite DNA instability and loss of heterozygosity in bronchial asthma.

Genetic alterations, such as loss of heterozygosity (LOH) or microsatellite instability (MI), have been reported in both malignant and benign disorders. In order to identify loci of deoxyribonucleic acid (DNA) mutation in asthma, MI and LOH were studied in sputum cells. DNA was extracted from cells in the sputum and blood cells of 22 patients with moderate asthma. Cells were analysed for MI and LOH using 18 polymorphic markers on chromosome 5q, 6p, 11q, 14q. Microsatellite analysis was also performed in six healthy subjects. None of the healthy individuals exhibited any genetic alteration. Genetic alterations were found in 16 of 22 asthmatic patients (73%). In total, 12 (54.5%) patients exhibited LOH only, one (4.5%) MI only, while three showed both MI and LOH. The highest incidence of LOH and MI was found on chromosome 14q. Mean immunoglobulin E and blood eosinophil levels were significantly higher in asthmatics with three or more genetic alterations. A high incidence of genetic alterations in the deoxyribonucleic acid of the sputum cells was found in asthmatic patients. Further studies are needed to identify the role of loss of heterozygosity and microsatellite instability in the investigation of genetic susceptibility of asthma and thus, in its pathogenesis.