Inflammatory Mediators Are Increased in Leukocytes of IVS-I-6 (T→C) Homozygous β-Thalassemia Intermedia.

Abstract 4068

Poster Board III-1003

Thalassemia is characterized by chronic inflammatory state and inflammation may play an important role in the pathogenesis of this diseases. Endothelial adhesion molecules, reactive oxygen species (ROS), C reactive protein and cytokines were increased in thalassemia patients. In addition, adherence of red blood cells, neutrophils and platelets to extracellular matrix protein and neutrophils chemotaxis has been shown to be markedly enhanced in β-thalassemia intermedia (TI) patients. Although alterations in inflammatory biomarkers have been related previously, most studies have been carried out with patients either polytransfused, receiving hydroxyurea (HU) therapy or with β-thalassemia major. An investigation of the role of inflammatory mediators in TI may further contribute to the understanding the pathogenesis of the disease. The aim of this study was to determine plasma levels and leukocyte gene expressions of inflammatory mediators/cytokines in neutrophils and mononuclear cells (MC) of untransfused patients and those not on HU therapy TI IVS-I-6 (T→C) homozygous (n≤20) and healthy controls (n≤20). ELISA was used to determine cytokine production; survivin protein expression was determined by flow cytometry with survivin-specific antibodies and qRT-PCR analysis to examine gene expression of cytokines, the protective enzyme heme oxygenase-1 (HO-1) and BIRC-5 (survivin). TNF-α, IL-6, IL-8 and IL-1β plasma levels were significantly higher in the plasma of TI individuals, when compared to control individuals (3.78 ± 0.4 vs 2.18 ± 0.3; 0.93 ± 0.1 vs 0.46 ± 0.08; 18.97 ± 5.6 vs 6.09 ± 1.1; 1.88 ± 0.7 vs 0.34 ± 0.05, pg/ml, P<0.05, respectively). Survivin protein levels in MC from TI was significantly increased when compared to healthy controls (45.9 ± 1.8 × 39.74 ± 1.74, MFI, P=0.022, respectively). IL-8, IL-10 and HO-1 gene expressions were unaltered in TI neutrophils (0.31 ± 0.13; 0.1 ± 0.08; 0.44 ± 0.2; A.U., respectively) compared to healthy controls neutrophils (0.45 ± 0.13; 0.17 ± 0.04; 0.22 ± 0.07, A.U., respectively), however in TI neutrophils gene expression of TNF-α was significantly higher than those of control subjects (0.76 ± 0.3 vs 0.24 ± 0.05, A.U., P=0.03, respectively). Expressions of genes encoding IL-8, IL-10, HO-1 and BIRC-5 were higher in MC of TI patients, compared with those of healthy controls (0.75 ± 0.3 vs 0.02 ± 0.005; 0.36 ± 0.07 vs 0.19 ± 0.04; 1.69 ± 0.28 vs 0.27 ± 0.09; 1.13 ± 0.12 vs 0.55 ± 0.17, A.U., P=0.02; P=0.04; P=0.0002, P=0.019, respectively). No significant alterations in the TNF-α mRNA levels were found in the MC of TI patients compared with healthy controls (0.21 ± 0.06 vs 0.16 ± 0.03, A.U., respectively). Taken together our data showed a higher production of inflammatory cytokines in MC, which could contribute to the pathophysiology of TI. Anti-inflammatory mechanisms (IL-10 and HO-1) were also up-regulated in these individuals, indicating efforts to counteract these alterations. These results are strong indications of the presence of chronic inflammatory processes and knowledge of these pathways may contribute to the understanding of the pathophysiology of TI.