Decreased TIM-3 Expression Level of Peripheral Blood Natural Killer Cells in Severe Aplastic Anemia

Severe aplastic anemia (SAA) is a life threatening disease characterized by severe pancytopenia and bone marrow failure. Natural killer (NK) cells are large granular lymphocytes which are one important component of the innate immune system and play a core role in regulation of adaptive immunity. T cell immunoglobulin mucin-3 (TIM-3), a member of the TIM family, appears to play an important negative regulatory role in T cells initial. Now, TIM-3 is widely detected on NK cells, and may contribute as a marker for activation and maturation of NK cells. Our previous studies have confirmed that the decrease of total NK cells, and CD56^bright, CD56^dim NK cell subsets and the higher expressions of NKp46 and perforin on NK cells may cause the over-function of T lymphocytes and thus lead to hematopoiesis failure in SAA. But, the expression of TIM-3 on NK cells in patients with SAA was still unknown.

In this study, we investigated the expression of TIM-3 and its mRNA on NK cells in peripheral blood of untreated and recovered SAA patients by flow cytometry and Real-time PCR. Results showed that the expression of TIM-3 on peripheral blood NK cells in SAA patients before IST was (63.57±12.14) %, which was significantly decreased than that in normal controls (85.62±9.03) % ( p<0.01). The expression of TIM-3 on CD56^dim NK cells was (66.41±11.74) % and (83.83±1.59) % separately in SAA patients before IST and normal controls. TIM-3 expressed in SAA patients before IST was lower than that in normal controls ( p<0.01). We also measured TIM-3 expressed on the surface of CD56^bright NK cells, but the result showed that there was no statistical difference between SAA patients before IST (61.11±24.99%) and normal controls (62.64±12.06%) (p>0.05). More interesting, the expression of TIM-3 on NK cells was (75.88±12.83) % in SAA patients after IST, which was significantly increased than that in SAA patients before IST, and was no difference with normal controls. As well, we found TIM-3 expression on both CD56^dim NK cells and CD56^bright NK cells in SAA patients after IST also has a rising trend compared with SAA patients before IST. However, these differences had no statistical significance. Further, we detected TIM-3 mRNA expression in NK cells isolated from peripheral blood. TIM-3 mRNA expression in peripheral blood NK cells from newly diagnosis SAA patients, recovering SAA patients and normal controls was evaluated, respectively. The relative TIM-3 mRNA expression was significantly increased in NK cells in SAA patients after IST compared with SAA patients before IST and normal controls, and this difference had statistical significance. Meanwhile, the relative TIM-3 mRNA expression was lower in NK cells in SAA patients before IST compared with normal controls. However, the difference had no statistical significance. In SAA patients, the expression of TIM-3 on NK cells was positively correlated with the level of WBC (r=0.685, p=0.000), proportion of neutrophil (r=0.825, p=0.000), and proportion of reticulocyte in peripheral blood (r=0.465, p=0.029). And it was negatively correlated with the proportion of lymphocyte in peripheral blood (r=-0.802, p=0.000).

According to our series studies, we hypothesize that the lower numbers and dysfunction of NK cells induce the failure of suppressing the over function of DC cells and T cells, that lead damage to healthy hematopoietic stem cells and the onset of SAA.