Search

The competency of antigen specific (Ag+) lymphocytes plays a critical role in the outcome of an adaptive immune response during an infection. Although the precursor frequency of Ag+ T cells often serves as an indicator, the evaluation of the proliferative capacity of Ag+ T cells of healthy humans to known viral antigens is limited. Here, we compared CD8 T cell responses to the antigen from cytomegalovirus (CMV-pp65) or influenza virus (Flu-M1) using an artificial antigen presenting system in vitro. We found that CD8 T cell responses (proliferation and Granzyme B production) to these two antigens varied in 23 healthy adults analyzed. The antigen specific CD8 T cell expansion did not correlate with the precursor frequency of total Ag+ CD8 T cells. Further comparing the response of isolated naïve (Tn) and central memory (Tcm) CD8 T cells showed that Tcm expanded more than Tn did to both antigens. Finally, we found that longer telomere lengths in CD8 T cells were positively correlated with the better expansion of these Ag+ CD8 T cells. Together, our study shows that antigen specific CD8 T cell response to CMV-pp65 and Flu-M1 is varied in healthy adults and the capacity of proliferation of these Ag+ CD8 T cells depends partly on the telomere length in these cells.

Genes containing both histone methylation H3K4me3 and H3K27me3 markers at the promoter are so called bivalent genes that have been implicated in stem cell differentiation and function. Our previous study identified several dozen bivalent genes in memory CD8 T cells (central memory, TCM and effector memory, TEM). But the role of bivalent genes in CD8 T cells is not understood. Our current focus is to determine the characteristics and functions of these bivalent genes during CD8 memory T cell activation. We used freshly isolated TCM and TEM from human blood that were stimulated by anti-CD3 plus anti-CD28 in vitro, and analyzed histone methylation and mRNA levels of these bivalent genes in resting and activated CD8 memory T cells. Our preliminary results showed that the bivalent status of most genes was stable but corresponding mRNA change was more detectable. We also identified a few bivalent genes whose status changed to become “monovalent”; either H3K4me3 or H3K27me3 after activation. We plan to determine the functional role of these bivalent genes that are altered after activation, and whether the bivalent histone methylation is present on a single gene allele or on two different alleles by sequential ChIP. Our study will shed new insight into the details of histone methylation on gene loci and the function of these bivalent genes in memory CD8 T cells. “This research was supported entirely by the Intramural Research Program of the NIH, National Institute on Aging “

The competency of antigen specific (Ag+) lymphocytes plays a critical role in the adaptive immune response. However, little is known about the differences in the competency of Ag+ T cells against different viruses. Here, we compared the CD8 T cell responses of 23 healthy adults to CMV (pp65) or Flu (M1) peptides using an in vitro artificial antigen presenting system. We found that CD8 T cell responses to CMV or Flu varied in both proliferation and Granzyme B production and were independent of the frequency of initial CMV+ or Flu+ CD8 T cells. In the comparison of the responses of Naïve (Tn) and central memory (Tcm) CD8 T cells, Tcm exhibited a greater expansion than did Tn for both CMV and Flu antigen. Intriguingly, the variations of the expansion between CMV+ and Flu+ CD8 T cells were observed only in Tcm but not Tn cells, indicating the importance of antigen experience. Furthermore, the expansion ability was correlated with the telomere length in the initial CD8 T cells positively and expanded Ag+ CD8 T cells negatively, suggesting a role for telomere length in cell proliferation. Taken together, this study showed that incompetency of Ag+ CD8 T cells is associated with antigen experience and short telomere length in healthy adults. Currently, we are analyzing the usage of the TCR variable genes (alpha and beta) of CMV+ or Flu+ CD8 T cells by a single cell RT-PCR-Sequencing method to gain additional information about the structure of these TCR against CMV and Flu antigens.

Krüppel-like factor 4 (Klf4), a zinc finger-containing transcription factor of the Krüppel-like factor family, plays critical role in cell differentiation, growth, and cycle. We report here a role of Klf4 in T cell development and differentiation. The expression of Klf4 was high in thymocytes and resting mature T cells, and was down-regulated in activated mature T cells. In the absence of Klf4, modest but significant reductions of T cell numbers in thymus (~27%) and in spleen (~25%), and a similar degree reduction of spleen CD4 T cells and a less degree loss of spleen CD8 T cells were found in Klf4 deficient mice. Gene expression analysis revealed an alteration of genes involved in cell cycling and proliferation in the absence of Klf4 in thymocytes, implying their role in control of T cell number. Furthermore, we found that significant reduction of IL-17 expressing CD4 T cells (TH-17) during in vitro differentiation in mice absence of Klf4 compared to the wild-type mice. We then demonstrated that Klf4 directly binds the promoter of IL17a but not to the promoter of ROR-γt gene, suggesting that Klf4 directly regulates Il17a in TH-17 differentiation. Together, these findings demonstrate that Klf4 regulates T cell development and TH-17 differentiation.