Detection of 24 TCR Vβ-Dβ1 sjTRECs in T Cells from Cord Blood, Peripheral Blood of Normal Individuals and Patients with AML-M2.

Thymic function is characterized its importance of thymus to T-cell diversity in the periphery of both children and adults during both health and disease. The generation of TCR diversity occurs in the thymus through recombination of gene segments encoding the variable parts of the TCRα and β chains. During these processes, by-products of the rearrangements are generated in the form of signal joint T-cell receptor excision circles (sjTRECs). As sjTRECs are stable extrachromosomal DNA fragments, are not replicated during mitosis and thus diluted with each round of cell division, and are therefore most frequent in naïve T cells that have recently left the thymus, their quantification is actually considered as a very valuable tool to estimate thymic function. Quantitative of δRec-ψJα sjTRECs can direct evaluate the recent thymic output function, but it is unable to analyze the particular thymic output function of different TCR Vβ subfamily naive T cells. The complexity of TCR Vβ repertoire is an important factor for immune reconstitution, it should be established the quantitative analysis of series TCR Vβ-Dβ sjTRECs to evaluate the levels of different Vβ subfamily naive T cells. In the present study, analysis of 24 TCR Vβ-Dβ sjTRECs was established by semi-nested PCR using 24 Vβ subfamily antisense primers and 2 Dβ1 sense primers. TCR Vβ-Dβ sjTRECs were amplified in genomic DNA from mononuclear cells of 10 cord blood samples, 10 cases of peripheral blood from normol individuals and 11 cases with AML-M2. Different amounts of DNA (corresponding to 2*10⁵, 5*10⁴, 1*10⁴ and 1*10³ cells respectively) from all samples were amplified to estimate the frequency of TCR Vβ-Dβ sjTRECs. The results showed that most Vβ subfamily sjTRECs could be detected in all samples from cord blood and peripheral blood at 2*10⁵ or 5*10⁴ cells level, some of Vβ subfamily sjTRECs could be detected in 1*10³ cells level. Whereas the frequency of Vβ subfamily sjTRECs were lower in peripheral blood T cells from patients with AML-M2 than in normal individuals (p<0.05). Vβ19, Vβ23 and Vβ24 subfamily sjTRECs could not be detected in all samples at 5*10⁴ cells level. The results indicated that Vβ subfamily naive T cells could be detected with different frequency in peripheral blood of normaol individuals as well as in some patients with AML-M2. Lower frequency of Vβ subfamily naive T cells was found in most AML-M2 patients.