Defective NATURAL Killer ACTIVITY Improved by IL-2 Stimulation Independent of CD16 and NKG2D Receptors IN PATIENTS with THALASSEMIA MAJOR.

Natural killer (NK) cells account for up to 15% of blood lymphocytes. Functional NK cells are found in the spleen. The function of NK cells is to recognize and kill virus infected cells and certain tumor cells. The mechanism of recognition is not fully understood, but it involves both activating and inhibitory receptors. CD 16 and NKG2D are receptors involved in two of the activation pathways of NK cells. NK cells use a variety of different mechanism to kill their targets. These include direct cell-cell signalling via surface molecules, and indirect signalling via cytokins. A decreased level of natural killer (NK) activity is one of the various immunological abnormalities in patients with thalassemia major.

This study aims to evaluate the relationship between NK cytotoxicity and CD 16 and NKG2D receptors, and effect of IL-2 stimulation in patients with thalassemia major.

The study includes 26 patients with thalassemia major (TM) with age ranged 7-35 (median age was 18, 17 of them were female and 9 male), and sex and age matched 16 normal healthy subjects as controls. Fourteen of the patients were splenectomized. Patients regulary chelated with deferasirox (n:16), deferipron (n: 8) and desferroxamin (n: 2). NK cells were isolated from PBMC of subjects by using RosetteSep. Before and after the IL2 incubation; CD16, NKG2D receptors are examined in NK cells of both b-thalassemia major patients and controls by flow cytometry and we analyzed cytolytic function of NK cells against K562 cells. The cytotoxic activity of NK cells was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Ratios of NK cells /K562 cells (Efector:Target (E:T)) were 1:1, 10:1 and 20:1.

NK cytotoxicity was found to be lower at E:T ratios of 1:1 and 10:1 in patients with thalassemia major when compared to controls(p < 0.05). Patients with above 18 years old had significantly lower NK activity than below 18 years old. Sex, presence of splenectomy and chelation type did not effect on NK activity (p>0.05). NK cytotoxicity significantly increased by IL-2 stimulation in 26 patients at E:T ratio of 10:1 (p<0.05). But the increase was still less than the control. There was no significantly statistically difference between CD16 and NKG2D receptor levels compared with patients and controls. Patient's CD 16 receptor levels did not change after incubation with IL-2. NKG2D receptor levels of control NK cells significantly increased after IL-2 incubation (2021.66 / 1675.79/molecule, p<0.003). We detected significantly increased levels of NKG2D after IL-2 incubation in patients with splenectomy than patients without splenectomy (1834.80 / 901.41/molecule, p<0.005).

IL-2 may have potential therapeutic effect to improve the defective NK activity in patients with thalassaemia major. Increasing effect of IL-2 stimulation on NK activity may be independent than NKG2D receptor pattern. Defective NK cell functions can be partially restored by interleukin IL-2.

This study was supported by Istanbul University Research Fund.

No relevant conflicts of interest to declare.