Soluble endothelial adhesion molecules and inflammation markers in patients with β-thalassemia intermedia

doi:10.1016/j.bcmd.2009.06.002

Blood Cells, Molecules, and Diseases

Volume 43, Issue 3, November–December 2009, Pages 230-234

https://doi.org/10.1016/j.bcmd.2009.06.002 Get rights and content

Abstract

The term thalassemia intermedia, indicates a clinical condition of intermediate severity between thalassaemia minor, the asymptomatic carrier, and thalassaemia major, the transfusion-dependent, severe form. Thromboembolic events frequently complicate the outcome of thalassemia intermedia patients, reflecting a hypercoagulable state to which endothelial activation is believed to play an important role. The aim of this study was to evaluate the levels of soluble endothelial adhesion molecules that reflect endothelial activation and dysfunction and levels of chronic inflammation markers in the serum of β-thalassemia intermedia patients. Thirty-five Greek patients with β-thalassemia intermedia that have received different types of treatment (Hydroxyurea, splenectomy, untreated), aged 8–63 years, were included in the study. Twenty apparently healthy individuals matched for age and sex, formed the control group. Measurements of sVCAM-1, sICAM-1, sTM, P-selectin, E-selectin and CRP levels were performed using immunoassays. We found that all endothelial adhesion molecules and CRP were significantly increased in patients (p < 0.001) and not influenced by treatment. A negative correlation was observed between levels of sICAM-1 and sTM and this finding agrees with the results of studies, which propose this correlation as a predictive marker of increased risk for vascular damage. No correlation was observed between endothelial adhesion molecules and inflammation markers. These findings support the hypothesis that a serious degree of endothelial activation and damage along with a state of chronic inflammation underlie the pathophysiology of β-thalassemia intermedia. Furthermore, these findings are of particular importance in patients who can otherwise be characterized by a subtle clinical phenotype and may have an important role in their clinical care.

Introduction

The term thalassemia intermedia is used to define a group of patients with β thalassemia in whom the clinical severity of the disease is somewhere between the mild symptoms of the β thalassemia trait and the severe manifestations of β thalassemia major. The diagnosis is a clinical one that is based on the patient maintaining a satisfactory hemoglobin (Hb) level of at least 6–7 g/dL at the time of diagnosis without the need for regular blood transfusions. This initial definition of thalassemia intermedia, which was based on clinical observation alone, retained its validity even after some of the specific mutations associated with thalassemia intermedia were recognized because severity of the clinical course remains unpredictable even in known genotypes. For this reason, some patients with a β-thalassemia intermedia genotype are treated as if they have thalassemia major because they present with severe manifestations; similarly, others with a thalassemia intermedia genotype are considered to have thalassemia minor because of the mild or even asymptomatic nature of their condition. This variability is most likely related to the presence or absence of modifying genes. Because of the significant overlap in clinical severity among the 3 types of β thalassemia and despite the fact that several genotypes are associated with the β thalassemia intermedia picture, the diagnosis continues to be a clinical one, regardless of the genotype involved [1]. Complications in thalassemia intermedia result from chronic hemolytic anemia, ineffective erythropoiesis and iron overload [2], [3]. Therapeutic approaches, such as iron chelation therapies, hydroxyurea and blood transfusions, have significantly increased the life expectancy of thalassemic patients, leading to the identification and description of previously underestimated conditions which complicate the life of such patients, such as thromboembolic phenomena [4].

Thromboembolic events are frequently described in thalassemia intermedia patients, clinically manifested as ischemic cerebral lesions, deep venous thrombosis and pulmonary hypertension [5], [6]. Autopsy findings confirm the presence of a high incidence of thromboembolic events [4] and Magnetic Resonance Imaging (MRI) studies have shown brain damage in otherwise asymptomatic thalassemic patients [7]. The above mentioned clinical, imaging and autopsy evidence define thalassemia as a hypercoagulable state [5]. Thalassemia intermedia patients suffer more frequently from thromboembolic events than patients with thalassemia major [4]. Pathogenesis of hypercoagulability in thalassemia is based on several factors, such as erythrocyte cell membrane alterations, platelet and endothelial cells activation [4].

Therapeutic approaches like splenectomy and transfusions seem to influence the manifestation of thromboembolic complications. Thus, splenectomized patients present higher risks for thrombosis than non-splenectomized patients, due probably to the presence of increased numbers of damaged erythrocytes as well as high thrombocyte counts [4], [8]. Regularly transfused patients present a lower incidence of thromboembolic phenomena than transfusion independent patients [8].

Inflammation is known to have an important role in the pathogenesis of thalassemia and a chronic inflammatory state is present in these patients. Pro-inflammatory cytokines such as IL-6 and inflammation markers such as CRP are known to be elevated in thalassemic patients [9], [10]. Endothelial activation is also believed to play an important role in the pathophysiology of thalassemia, through inflammation and thrombosis [4], [9], [11]. Endothelial activation is reflected by the increased serum levels of soluble endothelial adhesion molecules such as inter-cellular adhesion molecule 1 (sICAM-1), vascular cell adhesion molecule 1 (sVCAM-1), E-selectin (E-sel) and von Willebrand factor (vWF) [12], [13]. Increased levels of soluble thrombomodulin (sTM) have also been demonstrated in α and β thalassemia patients, indicating endothelial injury [14].

The aim of this study was to evaluate the levels of soluble endothelial adhesion molecules, that are used as markers of endothelial activation and dysfunction, in the serum of β-thalassemia intermedia patients. Levels of CRP as chronic inflammation marker were also evaluated.

Section snippets

Patients and methods

Thirty-five patients with thalassemia intermedia, 13 men and 22 women, aged 8–63 years, selected from the Laboratory of Human Genetics of Athens University and the First Department of Internal Medicine of the Medical School of Athens, were included in the study. The blood samples were collected in an outpatient basis, as patient's clinically steady state did not require hospitalization.

Seven (7/35) patients were smokers, while one (1/35) presented cardiac insufficiency, two (2/35) presented

Results

We compared serum levels of soluble endothelial molecules in β-thalassemia intermedia patients with healthy controls. As shown in Fig. 1, levels of sICAM-1 (p < 0.0001), sVCAM-1 (p < 0.0001), P-selectins (p < 0.0001) and E-selectins (p < 0.0001) are significantly higher compared to controls. Levels of sTM are also higher but results do not reach statistical significance (p = 0.65).

Patients were then divided in groups according to different therapeutic approaches. Eight patients (8/35) received

Discussion

Previous studies have shown higher levels of endothelial adhesion molecules, such as sICAM-1, sVCAM-1, E-sel and vWF, in the serum of patients with α and β thalassemia [12], [14]. In vitro studies came to the same conclusion [12]. In other clinical studies concerning transfusion-dependent thalassemia major patients, plasma levels of sICAM-1 and sVCAM-1 were found significantly increased [9], [13]. Plasma levels of sTM were also elevated in β-thalassemia patients and this team proposes the use

Acknowledgments

Grant/funding support: Funding were received from Athens University, (to I.P and J.R.). The funding sources played no role in the study design; in the collection, analysis, and interpretation of data; in the writing of the report; or in the decision to submit the report for publication.

Financial disclosures: None declared.

References (19)

A. Taher et al.
Thalassemia intermedia: revisited
Blood Cell. Mol. Dis.
(2006)
A.T. Taher et al.
Thalassemia and hypercoagulability
Blood Rev.
(2008)
A. Eldor et al.
The hypercoagulable state in thalassemia
Blood
(2002)
C. Aggeli et al.
Endothelial dysfunction and inflammatory process in transfusion-dependent patients with beta-thalassemia major
Int. J. Cardiol.
(2005)
C. Camaschella et al.
Thalassemia intermedia
Haematologica
(1995)
A. Aessopos et al.
Thalassemia intermedia today: should patients regularly receive transfusions?
Transfusion
(2007)
M. Karimi et al.
Cerebrovascular accident in beta-thalassemia major (beta-TM) and beta-thalassemia intermedia (beta-TI)
Am. J. Hematol.
(2008)
L. Manfre et al.
MR imaging of the brain: findings in asymptomatic patients with thalassemia intermedia and sickle cell-thalassemia disease
Am. J. Roentgenol.
(1999)
M.D. Cappellini et al.
Venous thromboembolism and hypercoagulability in splenectomized patients with thalassemia intermedia
Br. J. Haematol.
(2000)

There are more references available in the full text version of this article.

Cited by (32)

Hemopexin and albumin inhibit heme-induced macrophage activation while also enabling heme-LPS synergistic promotion of TNF production
2023, Advances in Redox Research
Free heme released from hemoglobin contributes to exacerbated inflammation and tissue damage in hemolytic diseases. While a moderate level of free heme does not cause intravascular inflammation by itself, its presence during infection greatly enhances inflammation. Although specific serum proteins have been found to affect heme-induced inflammation, the selective contribution of serum proteins inhibiting macrophage activation by heme or, conversely, amplifying the production of cytokines by macrophages stimulated with heme and microbial molecules, is poorly defined. Here we identified a serum fraction containing proteins with >50 KDa which was capable of inhibiting heme-stimulated TNF production and capable of enabling TNF production under conditions of a heme-LPS synergy. The inhibition of heme-induced TNF production was mimicked by Hemopexin (Hx), human serum albumin (HSA), serum from Hx-knockout mice, and less efficiently by serum from albumin-knockout mice, but not by serum LDL. Hx and HSA inhibited heme-induced ROS generation, MAPK/ Syk phosphorylation and cell death. However, Hx and HSA each also promoted the synergistic relationship between heme and LPS upon TNF production. Serum from Hx-knockout mice was fully capable of enabling this synergy, while serum from albumin-knockout mice was less efficient to promote TNF production under these conditions. Low concentrations of HSA mimicked the ability of serum to enable heme-stimulated IL-1β production after LPS priming, while high concentrations inhibited it. Together, our findings indicate how heme inflammatory effects are restrained in the blood upon sterile hemolysis, yet exacerbate inflammation in the presence of microbes. Moreover, it is interesting to note that opposing effects of serum proteins on heme-induced macrophage activation were selected through evolution, with both effects exerted by Hx and albumin.
Impaired bone marrow microenvironment and stem cells in transfusion-dependent beta-thalassemia
2022, Biomedicine and Pharmacotherapy
Citation Excerpt :
Circulating ADMA level has been shown to be significantly higher in children with BT major compared with those in a control group along with adhesion molecules slCAM-1, sVCAM-1, and P-selectin [61,62]. An increasing number of studies has also established that adhesion molecules (slCAM-1, sVCAM-1, E-selectin, P-selectin) and inflammatory factors (IL-6 and IL-1β) are at heightened levels in patients with thalassemia [63–66]. This is likely related to IO and stimulation of oxidative stress and macrophage activation.
Beta-thalassemia (BT) is a hereditary disease caused by abnormal hemoglobin synthesis with consequent ineffective erythropoiesis. Patients with thalassemia major are dependent on long-term blood transfusions with associated long-term complications such as iron overload (IO). This excess iron can result in tissue damage, impaired organ function, and increased morbidity. Growing evidence has demonstrated that IO contributes to impairment of the bone marrow (BM) microenvironment that largely impacts the function of BM mesenchymal stem cells, hematopoietic stem cells, and endothelial cells. In this article, we review recent progress in the understanding of iron metabolism and the perniciousness induced by IO. We highlight the importance of understanding the cross-talk between BM stem cells and the BM microenvironment, particularly the pathological effect of IO on BM stem cells and BT-associated complications. We also provide an update on recent novel therapies to cure transfusion-dependent beta-thalassemia and iron overload-induced complications for their future clinical application.
Vascular and hemostatic alterations associated with pulmonary hypertension in β-thalassemia hemoglobin E patients receiving regular transfusion and iron chelation
2019, Thrombosis Research
Citation Excerpt :
However, MP-TFA and TFAg levels were not significantly elevated suggesting hypercoagulability in β-thal/HbE with chronic transfusion was independent of circulating TF. Furthermore, sP-selectin levels that reflected platelet activation were not significantly increased compared with controls and did not differ between splenectomized and non-splenectomized subgroups as observed in previous reports which patients were mostly on non-regular transfusion [43,44]. Remarkably, TAT was the only prothrombotic variable significantly different between PH and non-PH subgroups and was significantly correlated with TRV.
Pulmonary hypertension (PH) is the commonest cardiac complication in β-thalassemia intermedia, including β-thalassemia/hemoglobin E (β-thal/HbE), and is strongly associated with splenectomy. We aimed to define the prevalence and comprehensively explore mechanisms of PH in β-thal/HbE patients receiving regular transfusion and iron chelation, which were reported to alleviate PH.
β-Thal/HbE patients receiving regular transfusion and iron chelation over one year were enrolled. Patients at risk for PH were defined by tricuspid-regurgitant-jet-velocity (TRV) ≥ 2.5 m/s. Laboratory and echocardiographic variables were compared with healthy controls.
There were 68 β-thal/HbE, including 31 (45.6%) splenectomized patients, and 38 controls included for analysis. PH was detected in 29 β-thal/HbE (42.6%). β-Thal/HbE with PH had a significant reduction in nitric oxide metabolites (NOx) but elevations in thrombin-antithrombin (TAT) complex, soluble thrombomodulin (sTM), endothelin-1 (ET-1) and flow-mediated dilation (FMD) values compared to those without PH (all, p < 0.05). TRV was significantly correlated with NOx, TAT, sTM, ET-1 and FMD values (r = −0.514, r = 0.281, r = 0.313, r = 0.245 and r = −0.474; all p < 0.05). Erythropoietic activity, serum ferritin, circulating total tissue factor (TF) antigen, microparticle-associated TF activity, microparticle's procoagulant activity and soluble p-selectin levels were not different between PH and non-PH subgroups. Notably, there were no significant associations between splenectomy and PH.
PH remains prevalent in β-thal/HbE patients receiving long-term transfusion and iron chelation. PH is not associated with splenectomy status but correlated with NO depletion, TF-independent hypercoagulability and endothelial perturbation.
Alterations of anticoagulant proteins and soluble endothelial protein C receptor in thalassemia patients of Chinese origin
2018, Thrombosis Research
Citation Excerpt :
As a marker of enhanced coagulation, TAT was significantly increased in this patient cohort and inversely correlated with AT III activity, which is in line with previous studies. ICAM-1, a known marker of endothelial activation and damage, was negatively correlated with TM which activates of PC and has vasoprotective activity [20]. The elevation of ICAM-1 observed in this study suggests that endothelial activation may be involved in thalassemia.
Thalassemia is characterized by a hypercoagulable state in which the protein C (PC) pathway controls thrombosis. We investigated changes in PC, protein S (PS), antithrombin III (AT III) and soluble endothelial protein C receptor (sEPCR) in thalassemia.
A group of 129 patients with β-thalassemia major (β-TM), β-thalassemia intermedia (β-TI), α-thalassemia intermedia (α-TI) and combined α-/β-thalassemia (α + β-thal) were compared with 32 gender- and age-matched controls. PC, PS, AT III, sEPCR, thrombin-antithrombin complex (TAT), and intercellular adhesion molecule1 (ICAM-1) antigens were measured by enzyme-linked immunosorbent assay. PC, AT III, and PS activity were assayed by substrate chromatography and a prothrombin time (PT)-based free protein S assay.
PC deficiency was seen in 95.3% of the patients and PS deficiency was seen in 77.5%. Concomitant reductions in PC and AT III antigen and activity were observed in β-TM, β-TI, and α-TI than in controls (p < 0.005). PC activity was lower in β-TM than in α-TI (p = 0.004). PS antigen was elevated in β-TM (p = 0.011) and sEPCR was elevated in α-TI (p = 0.018). Nonsplenectomized patients had lower PC (p = 0.001) and PS (p = 0.006) and higher sEPCR (p = 0.021) than postsplenectomy patients. Transfusion dependent thalassemia (TDT) patients had lower PC levels (p < 0.005) than those with nontransfusion dependent thalassemia (NTDT). ICAM-1 was increased in patient subgroups (p < 0.001), especially those with splenectomies (p = 0.009), and TAT was increased in all patient subgroups compared with controls (p < 0.001) except for α + β-thal.
Deficiencies of anticoagulant proteins and elevated sEPCR contributed to chronic hypercoagulability in these thalassemia patients of Chinese origin. Splenectomy alleviated these alterations in this patient cohort with the median duration since splenectomy of two years. Blood transfusion was not ideal for avoiding thrombosis.
The association between four SNPs (rs7482144, rs4671393, rs28384513 and rs4895441) and fetal hemoglobin levels in Chinese Zhuang β-thalassemia intermedia patients
2017, Blood Cells, Molecules, and Diseases
Citation Excerpt :
Previous study found that the HMIP was an important regulator of γ-globin gene in mice through modulating the MYB expression [31]. In addition, multiple cytokines and transcription factors motifs involved in γ-globin expression have also been identified to have negative or positive association with HbF level [32,33]. HBS1L, a GTPases, could influence HbF expression through indirectly effect on these factors [10].
Four SNPs (rs7482144, rs4671393, rs28384513 and rs4895441) associated with HbF levels have been identified in different populations worldwide. To explore whether these SNPs modulate HbF expression in Chinese Zhuang population, 436 Chinese Zhuang β-thalassemia intermedia (β-TI) patients were divided into high HbF level group (mean HbF = 25.5%, n = 218) and low group (mean HbF = 6.51%, n = 218) for genotyping using PCR-HRM method. Results demonstrated that there was a significantly higher minor allele frequency (MAF = 34.2%) of rs4895441 (G) in HMIP in high HbF level group than that in low group (MAF = 19.8%) (P = 0.001, OR = 1.73, 95% CI: 1.24–2.57). The cumulative effects of risk genotypes of these loci for patients carrying any combination of 1, 2 or 3 risk genotype had a gradually increased risk of high HbF level phenotype compared to those without the risk genotypes (OR = 1.50–9.06, P = 0.0008); Gene-gene interaction of rs7842144 and rs4895441 showed the best model with the smallest prediction error (0.4259) and the greatest consistency of coefficient of variation (P = 0.01). We concluded that rs4895441, G on HMIP might be a high-risk modifier variant for high HbF level expression, and HBG2, BCL11A and HMIP genes, as HbF quantitative trait loci (QTL) could have a synergistic effect on increasing the HbF level in Chinese Zhuang β-TI patients.
Plasma neutrophil gelatinase-associated lipocalin levels are markedly increased in patients with non-transfusion-dependent thalassemia: Lack of association with markers of erythropoiesis, iron metabolism and renal function
2014, Clinical Biochemistry
Citation Excerpt :
Interestingly, we found that NGAL levels correlated with hs-CRP levels. We have recently reported that patients with TI as well as patients with sickle cell-beta thalassemia have a chronic low-grade inflammation [20,21]. Low-grade inflammation has been implicated in the etiology of clusters of so-called metabolic syndromes, such as insulin resistance, visceral adiposity and atherosclerosis.
Neutrophil Gelatinase-Associated Lipocalin (NGAL) (known as NGAL, Lipocalin 2, Siderocalin, Uterocalin, proteinase-3 and 24p3) is a mammalian small 25-kD peptide that belongs to the lipocalin superfamily, which consists of about 20 small lipoproteins. NGAL was initially discovered as an antibacterial factor of natural immunity and an acute-phase protein. NGAL is also an iron trafficking protein, a member of the non-transferrin-bound iron (NTBI) pool and an alternative to the transferrin-mediated iron-delivery pathway. Of note, NTBI, which is elevated in thalassemic patients, induces cellular toxicity. In this study we investigated the possible association of NGAL with parameters of erythropoiesis, iron metabolism and renal injury in patients with non-transfusion-dependent thalassemia (thalassemia intermedia or TI).
Thirty-five patients with TI, 13 men and 22 women, aged 8–63 years, were included in the study, while, 20 healthy individuals served as controls. Plasma NGAL levels were determined using an immunoenzymatic technique. Erythroid marrow activity was estimated by measuring soluble transferrin receptors (sTfR) levels with a turbidimetric technique. NTBI levels were determined using electrothermal atomic absorption spectrometry. Cystatin C, β2-microglobulin and hs-CRP concentrations were measured by means of immunonephelometric techniques.
The main results of the study showed: a) NGAL levels were significantly higher in patients with TI compared to controls (139.1 ± 86.1 vs 51.2 ± 11.8 μg/L, p < 0.0001), without significant effect of splenectomy or hydroxyurea on NGAL levels. Only 4 patients had NGAL levels within the control group range, b) no correlation was found between NGAL levels and either the parameters of erythropoiesis Hb, Hb F, reticulocytes and sTfR (p > 0.66, p > 0.67, p > 0.63 and p > 0.81 respectively), or with those of iron metabolism ferritin and NTBI (p > 0.90 and p > 0.95 respectively).
The increased NGAL levels reported for the first time in TI patients in this study are in agreement with the elevated expression of NGAL observed in TI mouse models. We postulate that the induction of NGAL in these patients may represent either a survival response, facilitating the survival of the less damaged thalassemic erythroid precursors, or a consequence of the abnormal iron regulation in TI.

View all citing articles on Scopus

View full text

Soluble endothelial adhesion molecules and inflammation markers in patients with β-thalassemia intermedia

Abstract

Introduction

Section snippets

Patients and methods

Results

Discussion

Acknowledgments

Blood Cell. Mol. Dis.

Blood Rev.

Blood

Int. J. Cardiol.

Thalassemia intermedia

Haematologica

Thalassemia intermedia today: should patients regularly receive transfusions?

Transfusion

Cerebrovascular accident in beta-thalassemia major (beta-TM) and beta-thalassemia intermedia (beta-TI)

Am. J. Hematol.

MR imaging of the brain: findings in asymptomatic patients with thalassemia intermedia and sickle cell-thalassemia disease

Am. J. Roentgenol.

Venous thromboembolism and hypercoagulability in splenectomized patients with thalassemia intermedia

Br. J. Haematol.