Abnormal Expression Of Shelterin Is Associated With Short Telomere and Immune Disorder In Severe Aplastic Anemia

To identify changes in telomere length and gene expression of shelterin (which is composed of six core components: TRF1, TRF2, POT1, TIN2, TPP1 and RAP1) in severe aplastic anemia(SAA). Looking for the reason why telomere getting short in SAA patients and the pathway that abnormal telomeres cause AA.

15 controls and 27 cases of SAA were enrolled in this study. There were 9 untreated and 18 recovering patients. Peripheral white blood cells(PWBCs)’ telomere length was tested by southern blot and gene expression of shelterin were detected by RT-PCR. Bone marrow mononuclear cells (BMMNC) from controls were cultured in different culture medium which were fetal calf serum only , SAA serum only, fetal calf serum containing TNF-α and fetal calf serum containing IFN-γ respectively. The apoptosis of these cells were detected by flowcytometry. The mRNA expression of POT1 were tested by RT-PCR , all the protein below including ATR, phosphorylated ATR, and phosphorylated ATM/ATR substrate were detected by western blot.

Telomeres length of PWBCs were found significantly shorter in SAA untreated group (4.89±1.66)kb and recovering group (7.04± 1.47 )kb than control group (11.65±5.55)kb (P<0.05). Telomeres Length of PWBCs turned shortening in accordance with T_H/S (CD4+Tcells/CD8+Tcells ratio) decreasing (r=0.593, p=0.007). The mRNA expression of POT1 decreased in SAA untreated group( 0.29(0.09-0.58)) than in recovering group( 0.78(0.22-2.69)) and controls (1.88(0.93-3.81)), (P<0.05). No differences about mRNA expression of TRF1、TRF2、TIN2、TPP1 and RAP1 were found among SAA untreated group, recovering group and controls in PWBCs. Undoubtedly, cells that cultured in SAA serum got more apoptosis(13.88±4.55) than that cultured in fetal calf serum(9.85±2.67)(P<0.05). The mRNA expression of POT1 was significantly declined in cells that from SAA patient uncultured group (0.45±0.29) , normal BMMNCs cultured by AA serum group (0.59±0.49) and IFN-γ group (0.40±0.35) than the cells cultured only by fetal calf serum group (1.19±0.62). ATR, phosphorylated ATR, and phosphorylated ATM/ATR substrate were found significantly increase in cells from AA serum group , TNF-α group and IFN-γ group than fetal calf serum group.

SAA patients have short PWBCs’ telomeres length which was in accordance with T_H/S.TNF-α and IFN-γ which were found at high concentration in SAA decades before may be the effective component that can trigger apoptosis through POT1 and the pathway of ATM/ATR and eventually lead to SAA.

No relevant conflicts of interest to declare.